Joseph (Joe) Lipsick

All Publications

• A History of Cancer Research: Oncogenic Transcription Factors. Cold Spring Harbor perspectives in biology Lipsick, J. 2024; 16 (6)

  Abstract

  Transcription factors play crucial roles in cancer, and oncogenic counterparts of cellular transcription factors are present in a number of tumor viruses. It was studies in the early 1980s that first showed tumor viruses could encode nuclear as well as cytoplasmic oncoproteins. Subsequent work provided detailed insight into their mechanisms of action, as well as potential therapeutic avenues. In this excerpt from his forthcoming book on the history of cancer research, Joe Lipsick looks back at early work on nuclear oncogenes, including the discovery of MYC, MYB, FOS and JUN, Rel/NF-κB, and nuclear receptors such as the retinoic acid receptor and thyroid hormone receptor.

  View details for DOI 10.1101/cshperspect.a035881

  View details for PubMedID 38830699

  View details for PubMedCentralID PMC11146308

• A History of Cancer Research: Retroviral Insertional Mutagenesis. Cold Spring Harbor perspectives in biology Lipsick, J. 2023; 15 (7)

  Abstract

  Early work on cancer showed that some retroviruses contain oncogenes that promote tumorigenesis, but how viruses that do not contain oncogenes could cause cancer was unclear. A series of studies in the 1980s uncovered another mechanism: insertional mutagenesis in which viral sequences drove aberrant expression of endogenous cellular proto-oncogenes. In this excerpt from his forthcoming book on the history of cancer research, Joe Lipsick looks back at these discoveries, how the work led to identification of new oncogenes and tumor suppressors, and the perils of the phenomenon for early gene therapy.

  View details for DOI 10.1101/cshperspect.a035873

  View details for PubMedID 37407069

• A History of Cancer Research: Retroviral Oncogenes. Cold Spring Harbor perspectives in medicine Lipsick, J. 2022; 12 (4)

  Abstract

  The discovery and characterization of retroviral oncogenes were important milestones in cancer research. The viruses turned out not to be key causes of cancer in humans, but the oncogenes they carried provided key clues to the role cellular genes, the proto-oncogene counterparts of these sequences, played in tumorigenesis. In this excerpt from his forthcoming book, Joe Lipsick looks back at early work on retroviruses, such as the experiments that distinguished their ability to infect and transform cells, the groundbreaking work on Src, and some of the controversy surrounding the Nobel Prize awarded for these discoveries.

  View details for DOI 10.1101/cshperspect.a035865

  View details for PubMedID 35581009

• A History of Cancer Research: Tumor Viruses. Cold Spring Harbor perspectives in biology Lipsick, J. 2021; 13 (6)

  Abstract

  Early studies of transmissible tumors in chickens provided evidence that viruses such as avian leukosis virus (ALV) and Rous sarcoma virus (RSV) can cause cancer in these animals. Doubts about the relevance to human tumors and failures to replicate some early work meant the field of tumor virology followed a bumpy course. Nevertheless, viruses that can cause cancers in rodents and humans were ultimately identified, and several Nobel prizes were awarded for work in this area. In this excerpt from his forthcoming book on the history of cancer research, Joe Lipsick looks back at the early history of tumor virus research, from some of the early false starts and debates, to discovery of reverse transcriptase, and identification of human papilloma virus (HPV) as the major cause of cervical cancer.

  View details for DOI 10.1101/cshperspect.a035774

  View details for PubMedID 34074674

• A History of Cancer Research: Carcinogens and Mutagens. Cold Spring Harbor perspectives in medicine Lipsick, J. 2021; 11 (3)

  Abstract

  Observations of the incidence of tumors among chimney sweeps in the eighteenth century and later experiments with coal tars provided early evidence that carcinogens in the environment can promote cancer. Subsequent studies of individuals exposed to radiation, work on fly genetics, and the discovery that DNA was the genetic material led to the idea that these carcinogens act by inducing mutations in DNA that change the behavior of cells and ultimately cause cancer. In this excerpt from his forthcoming book, Joe Lipsick looks back at how the concepts of mutagenesis and carcinogenesis emerged, how these converged with development of the Ames test, and how biochemistry and crystallography ultimately revealed the underlying molecular basis.

  View details for DOI 10.1101/cshperspect.a035857

  View details for PubMedID 33649023

• A History of Cancer Research: Tumor Suppressor Genes. Cold Spring Harbor perspectives in biology Lipsick, J. n. 2020; 12 (2)

  Abstract

  Tumor suppressor genes encode critical intracellular regulators, such as the retinoblastoma protein. They control processes including cell proliferation, cell survival, and responses to DNA damage and are frequently mutated in cancer. In this excerpt from his forthcoming book on the history of cancer research, Joe Lipsick looks back at the discovery of tumor suppressor genes, covering the early work on cell fusion by Henry Harris, Knudson's two-hit hypothesis, the genetic mapping studies that first identified the RB gene, and subsequent work on silencing.

  View details for DOI 10.1101/cshperspect.a035907

  View details for PubMedID 32015099

• A long lost key opens an ancient lock: Drosophila Myb causes a synthetic multivulval phenotype in nematodes. Biology open Vorster, P. J., Goetsch, P. n., Wijeratne, T. U., Guiley, K. Z., Andrejka, L. n., Tripathi, S. n., Larson, B. J., Rubin, S. M., Strome, S. n., Lipsick, J. S. 2020

  Abstract

  The five-protein MuvB core complex is highly conserved in animals. This nuclear complex interacts with RB family tumor suppressor proteins and E2F-DP transcription factors to form DREAM complexes that repress genes that regulate cell cycle progression and cell fate. The MuvB core complex also interacts with proteins Myb family oncoproteins to form the Myb-MuvB complexes that activate many of the same genes. We show that animal-type Myb genes are present in Bilateria, Cnidaria, and Placozoa, the latter including the simplest known animal species. However, bilaterian nematode worms lost their animal-type Myb genes hundreds of millions of years ago. Nevertheless, amino acids in the LIN9 and LIN52 proteins that directly interact with the MuvB-binding domains of human B-Myb and Drosophila Myb are conserved in C. elegans. Here we show that, despite greater than 500 million years since their last common ancestor, the Drosophila melanogaster Myb protein can bind to the nematode LIN9-LIN52 proteins in vitro and can cause a synthetic multivulval (synMuv) phenotype in vivo. This phenotype is similar to that caused by loss-of-function mutations in C. elegans synMuvB class genes including those that encode homologs of the MuvB core, RB, E2F, and DP. Furthermore, amino acid substitutions in the MuvB-binding domain of Drosophila Myb that disrupt its functions in vitro and in vivo also disrupt these activities in C. elegans. We speculate that nematodes and other animals may contain another protein that can bind to LIN9 and LIN52 in order to activate transcription of genes repressed by DREAM complexes.

  View details for DOI 10.1242/bio.051508

  View details for PubMedID 34004684

• A History of Cancer Research: Tyrosine Kinases. Cold Spring Harbor perspectives in biology Lipsick, J. 2019; 11 (2)

  Abstract

  Proteins containing tyrosine kinase activity play critical roles in cancer signaling. Intracellular SRC-family kinases relay growth signals from numerous cell surface receptors and can be constitutively activated by oncogenic mutations, as can transmembrane growth factor receptors such as epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER2) that signal via their tyrosine kinase activity. In this excerpt from his forthcoming book on the history of cancer research, Joe Lipsick looks back at the discovery of tyrosine kinases and the demonstration that the V-SRC protein encoded by Rous sarcoma virus was a tyrosine kinase.

  View details for PubMedID 30709881

• Structural mechanism of Myb-MuvB assembly PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Guiley, K. Z., Iness, A. N., Saini, S., Tripathi, S., Lipsick, J. S., Litovchick, L., Rubin, S. M. 2018; 115 (40): 10016–21

  Abstract

  The MuvB transcriptional regulatory complex, which controls cell-cycle-dependent gene expression, cooperates with B-Myb to activate genes required for the G2 and M phases of the cell cycle. We have identified the domain in B-Myb that is essential for the assembly of the Myb-MuvB (MMB) complex. We determined a crystal structure that reveals how this B-Myb domain binds MuvB through the adaptor protein LIN52 and the scaffold protein LIN9. The structure and biochemical analysis provide an understanding of how oncogenic B-Myb is recruited to regulate genes required for cell-cycle progression, and the MMB interface presents a potential therapeutic target to inhibit cancer cell proliferation.

  View details for PubMedID 30224471

• The role of variant histone H2AV in D. melanogaster larval hematopoiesis Development Grigorian, M., DeBruhl, H., Lipsick, J. S. 2017; 144 (8): 1441-1449

  View details for DOI 10.1242/dev.142729

• The Drosophila LIN54 homolog Mip120 controls two aspects of oogenesis. Biology open Cheng, M. H., Andrejka, L. n., Vorster, P. J., Hinman, A. n., Lipsick, J. S. 2017

  Abstract

  The conserved multi-protein MuvB core associates with the Myb oncoproteins and with the RB-E2F-DP tumor suppressor proteins in complexes that regulate cell proliferation, differentiation, and apoptosis. Drosophila Mip120, a homolog of LIN54, is a sequence-specific DNA-binding protein within the MuvB core. A mutant of Drosophila mip120 was previously shown to cause female and male sterility. We now show that Mip120 regulates two different aspects of oogenesis. First, in the absence of the Mip120 protein, egg chambers arrest during the transition from stage 7 to 8 with a failure of the normal program of chromosomal dynamics in the ovarian nurse cells. Specifically, the decondensation, disassembly and dispersion of the endoreplicated polytene chromosomes fail to occur without Mip120. The conserved carboxy-terminal DNA-binding and protein-protein interaction domains of Mip120 are necessary but are not sufficient for this process. Second, we show that a lack of Mip120 causes a dramatic increase in the expression of benign gonial cell neoplasm (bgcn), a gene that is normally expressed in only a small number of cells within the ovary including the germline stem cells.

  View details for PubMedID 28522430

• Recurrent rearrangements of the Myb/SANT-like DNA-binding domain containing 3 gene (MSANTD3) in salivary gland acinic cell carcinoma. PloS one Barasch, N., Gong, X., Kwei, K. A., Varma, S., Biscocho, J., Qu, K., Xiao, N., Lipsick, J. S., Pelham, R. J., West, R. B., Pollack, J. R. 2017; 12 (2)

  Abstract

  Pathogenic gene fusions have been identified in several histologic types of salivary gland neoplasia, but not previously in acinic cell carcinoma (AcCC). To discover novel gene fusions, we performed whole-transcriptome sequencing surveys of three AcCC archival cases. In one specimen we identified a novel HTN3-MSANTD3 gene fusion, and in another a novel PRB3-ZNF217 gene fusion. The structure of both fusions was consistent with the promoter of the 5' partner (HTN3 or PRB3), both highly expressed salivary gland genes, driving overexpression of full-length MSANTD3 or ZNF217. By fluorescence in situ hybridization of an expanded AcCC case series, we observed MSANTD3 rearrangements altogether in 3 of 20 evaluable cases (15%), but found no additional ZNF217 rearrangements. MSANTD3 encodes a previously uncharacterized Myb/SANT domain-containing protein. Immunohistochemical staining demonstrated diffuse nuclear MSANTD3 expression in 8 of 27 AcCC cases (30%), including the three cases with MSANTD3 rearrangement. MSANTD3 displayed heterogeneous expression in normal salivary ductal epithelium, as well as among other histologic types of salivary gland cancer though without evidence of translocation. In a broader survey, MSANTD3 showed variable expression across a wide range of normal and neoplastic human tissue specimens. In preliminary functional studies, engineered MSANTD3 overexpression in rodent salivary gland epithelial cells did not enhance cell proliferation, but led to significant upregulation of gene sets involved in protein synthesis. Our findings newly identify MSANTD3 rearrangement as a recurrent event in salivary gland AcCC, providing new insight into disease pathogenesis, and identifying a putative novel human oncogene.

  View details for DOI 10.1371/journal.pone.0171265

  View details for PubMedID 28212443

• The Complex Containing Drosophila Myb and RB/E2F2 Regulates Cytokinesis in a Histone H2Av-Dependent Manner MOLECULAR AND CELLULAR BIOLOGY Debruhl, H., Wen, H., Lipsick, J. S. 2013; 33 (9): 1809-1818

  Abstract

  In Drosophila, mutation of the oncogene Myb reduced the expression of mitotic genes, such as polo and ial, and caused multiple mitotic defects, including disrupted chromosome condensation and abnormal spindles. We now show that binucleate cells, the hallmark phenotype of cytokinesis failure, accumulate in Myb-null ovarian follicle cell and wing disc epithelia. Myb functions as an activator in the generally repressive Drosophila RBF, E2F2, and Myb (dREAM)/Myb-MuvB complex. Absence of the dREAM subunit Mip130 or E2F2 suppressed the Myb-null cytokinesis defect. Therefore, we used Myb-null binucleate cells as a quantitative phenotypic readout of transcriptional repression by the dREAM complex. In the absence of Myb, the complex was sensitive to the dose of the subunits E2F2, Mip120, Caf1, and Lin-52 but not Mip130 or Mip40. Surprisingly, reduction of the dose of His2Av/H2A.z also suppressed the Myb-null binucleate cell phenotype, suggesting a novel role for this variant histone in transcriptional repression by the dREAM complex.

  View details for DOI 10.1128/MCB.01401-12

  View details for Web of Science ID 000317273300011

  View details for PubMedID 23438598

  View details for PubMedCentralID PMC3624180

• Duplication and maintenance of the Myb genes of vertebrate animals BIOLOGY OPEN Davidson, C. J., Guthrie, E. E., Lipsick, J. S. 2013; 2 (2): 101-110

  Abstract

  Gene duplication is an important means of generating new genes. The major mechanisms by which duplicated genes are preserved in the face of purifying selection are thought to be neofunctionalization, subfunctionalization, and increased gene dosage. However, very few duplicated gene families in vertebrate species have been analyzed by functional tests in vivo. We have therefore examined the three vertebrate Myb genes (c-Myb, A-Myb, and B-Myb) by cytogenetic map analysis, by sequence analysis, and by ectopic expression in Drosophila. We provide evidence that the vertebrate Myb genes arose by two rounds of regional genomic duplication. We found that ubiquitous expression of c-Myb and A-Myb, but not of B-Myb or Drosophila Myb, was lethal in Drosophila. Expression of any of these genes during early larval eye development was well tolerated. However, expression of c-Myb and A-Myb, but not of B-Myb or Drosophila Myb, during late larval eye development caused drastic alterations in adult eye morphology. Mosaic analysis implied that this eye phenotype was cell-autonomous. Interestingly, some of the eye phenotypes caused by the retroviral v-Myb oncogene and the normal c-Myb proto-oncogene from which v-Myb arose were quite distinct. Finally, we found that post-translational modifications of c-Myb by the GSK-3 protein kinase and by the Ubc9 SUMO-conjugating enzyme that normally occur in vertebrate cells can modify the eye phenotype caused by c-Myb in Drosophila. These results support a model in which the three Myb genes of vertebrates arose by two sequential duplications. The first duplication was followed by a subfunctionalization of gene expression, then neofunctionalization of protein function to yield a c/A-Myb progenitor. The duplication of this progenitor was followed by subfunctionalization of gene expression to give rise to tissue-specific c-Myb and A-Myb genes.

  View details for DOI 10.1242/bio.20123152

  View details for Web of Science ID 000209205900001

  View details for PubMedCentralID PMC3575645

• Epigenetic regulation of olfactory receptor gene expression by the Myb-MuvB/dREAM complex GENES & DEVELOPMENT Sim, C. K., Perry, S., Tharadra, S. K., Lipsick, J. S., Ray, A. 2012; 26 (22): 2483-2498

  Abstract

  In both mammals and insects, an olfactory neuron will usually select a single olfactory receptor and repress remaining members of large receptor families. Here we show that a conserved multiprotein complex, Myb-MuvB (MMB)/dREAM, plays an important role in mediating neuron-specific expression of the carbon dioxide (CO(2)) receptor genes (Gr63a/Gr21a) in Drosophila. Activity of Myb in the complex is required for expression of Gr63a/Gr21a and acts in opposition to the histone methyltransferase Su(var)3-9. Consistent with this, we observed repressive dimethylated H3K9 modifications at the receptor gene loci, suggesting a mechanism for silencing receptor gene expression. Conversely, other complex members, Mip120 (Myb-interacting protein 120) and E2F2, are required for repression of Gr63a in inappropriate neurons. Misexpression in mutants is accompanied by an increase in the H3K4me3 mark of active chromatin at the receptor gene locus. Nuclei of CO(2) receptor-expressing neurons contain reduced levels of the repressive subunit Mip120 compared with surrounding neurons and increased levels of Myb, suggesting that activity of the complex can be regulated in a cell-specific manner. Our evidence suggests a model in which olfactory receptors are regulated epigenetically and the MMB/dREAM complex plays a critical role in specifying, maintaining, and modulating the receptor-to-neuron map.

  View details for DOI 10.1101/gad.201665.112

  View details for Web of Science ID 000311275800006

  View details for PubMedID 23105004

  View details for PubMedCentralID PMC3505819

• Drosophila Lin-52 Acts in Opposition to Repressive Components of the Myb-MuvB/dREAM Complex MOLECULAR AND CELLULAR BIOLOGY Lewis, P. W., Sahoo, D., Geng, C., Bell, M., Lipsick, J. S., Botchan, M. R. 2012; 32 (16): 3218-3227

  Abstract

  The Drosophila melanogaster Myb-MuvB/dREAM complex (MMB/dREAM) participates in both the activation and repression of developmentally regulated genes and origins of DNA replication. Mutants in MMB subunits exhibit diverse phenotypes, including lethality, eye defects, reduced fecundity, and sterility. Here, we used P-element excision to generate mutations in lin-52, which encodes the smallest subunit of the MMB/dREAM complex. lin-52 is required for viability, as null mutants die prior to pupariation. The generation of somatic and germ line mutant clones indicates that lin-52 is required for adult eye development and for early embryogenesis via maternal effects. Interestingly, the maternal-effect embryonic lethality, larval lethality, and adult eye defects could be suppressed by mutations in other subunits of the MMB/dREAM complex. These results suggest that a partial MMB/dREAM complex is responsible for the lethality and eye defects of lin-52 mutants. Furthermore, these findings support a model in which the Lin-52 and Myb proteins counteract the repressive activities of the other members of the MMB/dREAM complex at specific genomic loci in a developmentally controlled manner.

  View details for DOI 10.1128/MCB.00432-12

  View details for Web of Science ID 000307203500003

  View details for PubMedID 22688510

  View details for PubMedCentralID PMC3434544

• Animal-specific C-terminal domain links myeloblastosis oncoprotein (Myb) to an ancient repressor complex PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Andrejka, L., Wen, H., Ashton, J., Grant, M., Iori, K., Wang, A., Manak, J. R., Lipsick, J. S. 2011; 108 (42): 17438-17443

  Abstract

  Members of the Myb oncoprotein and E2F-Rb tumor suppressor protein families are present within the same highly conserved multiprotein transcriptional repressor complex, named either as Myb and synthetic multivuval class B (Myb-MuvB) or as Drosophila Rb E2F and Myb-interacting proteins (dREAM). We now report that the animal-specific C terminus of Drosophila Myb but not the more highly conserved N-terminal DNA-binding domain is necessary and sufficient for (i) adult viability, (ii) proper localization to chromosomes in vivo, (iii) regulation of gene expression in vivo, and (iv) interaction with the highly conserved core of the MuvB/dREAM transcriptional repressor complex. In addition, we have identified a conserved peptide motif that is required for this interaction. Our results imply that an ancient function of Myb in regulating G2/M genes in both plants and animals appears to have been transferred from the DNA-binding domain to the animal-specific C-terminal domain. Increased expression of B-MYB/MYBL2, the human ortholog of Drosophila Myb, correlates with poor prognosis in human patients with breast cancer. Therefore, our results imply that the specific interaction of the C terminus of Myb with the MuvB/dREAM core complex may provide an attractive target for the development of cancer therapeutics.

  View details for DOI 10.1073/pnas.1111855108

  View details for Web of Science ID 000295975300045

  View details for PubMedID 21969598

  View details for PubMedCentralID PMC3198365

• MYB Expression and Translocation in Adenoid Cystic Carcinomas and Other Salivary Gland Tumors With Clinicopathologic Correlation AMERICAN JOURNAL OF SURGICAL PATHOLOGY West, R. B., Kong, C., Clarke, N., Gilks, T., Lipsick, J. S., Cao, H., Kwok, S., Montgomery, K. D., Varma, S., Le, Q. 2011; 35 (1): 92-99

  Abstract

  Adenoid cystic carcinoma is a locally aggressive salivary gland neoplasm, which has a poor long-term prognosis. A chromosomal translocation involving the genes encoding the transcription factors, MYB and NFIB, has been recently discovered in these tumors.MYB translocation and protein expression were studied in 37 adenoid cystic carcinomas, 112 other salivary gland neoplasms, and 409 nonsalivary gland neoplasms by fluorescence in situ hybridization and immunohistochemistry. MYB translocation and expression status in adenoid cystic carcinoma was correlated with clinicopathologic features including outcome, with a median follow-up of 77.1 months (range, 23.2 to 217.5 mo) for living patients.A balanced translocation between MYB and NFIB is present in 49% of adenoid cystic carcinomas but is not identified in other salivary gland tumors or nonsalivary gland neoplasms. There is no apparent translocation of MYB in 35% of the cases. Strong Myb immunostaining is very specific for adenoid cystic carcinomas but is only present in 65% of all cases. It is interesting to note that Myb immunostaining is confined to the basal cell component although the translocation is present in all the cells. Neoplasms with MYB translocation show a trend toward higher local relapse rates, but the results are not statistically significant with the current number of cases.MYB translocation and expression are useful diagnostic markers for a subset of adenoid cystic carcinomas. The presence of the translocation may be indicative of local aggressive behavior, but a larger cohort may be required to show statistical significance.

  View details for DOI 10.1097/PAS.0b013e3182002777

  View details for PubMedID 21164292

• The C-MYB story-is it definitive? PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Lipsick, J. S. 2010; 107 (40): 17067-17068

  View details for DOI 10.1073/pnas.1012402107

  View details for Web of Science ID 000282512000005

  View details for PubMedID 20858731

  View details for PubMedCentralID PMC2951432

• A conserved acidic patch in the Myb domain is required for activation of an endogenous target gene and for chromatin binding MOLECULAR CANCER Ko, E. R., Ko, D., Chen, C., Lipsick, J. S. 2008; 7

  Abstract

  The c-Myb protein is a transcriptional regulator initially identified by homology to the v-Myb oncoprotein, and has since been implicated in human cancer. The most highly conserved portion of the c-Myb protein is the DNA-binding domain which consists of three imperfect repeats. Many other proteins contain one or more Myb-related domains, including a number of proteins that do not bind directly to DNA. We performed a phylogenetic analysis of diverse classes of Myb-related domains and discovered a highly conserved patch of acidic residues common to all Myb-related domains. These acidic residues are positioned in the first of three alpha-helices within each of the three repeats that comprise the c-Myb DNA-binding domain. Interestingly, these conserved acidic residues are present on a surface of the protein which is distinct from that which binds to DNA. Alanine mutagenesis revealed that the acidic patch of the third c-Myb repeat is essential for transcriptional activity, but neither for nuclear localization nor DNA-binding. Instead, these acidic residues are required for efficient chromatin binding and interaction with the histone H4 N-terminal tail.

  View details for DOI 10.1186/1476-4598-7-77

  View details for Web of Science ID 000260394300001

  View details for PubMedID 18840288

  View details for PubMedCentralID PMC2572630

• Epigenetic regulation of gene expression by Drosophila Myb and E2F2-RBF via the Myb-MuvB/dREAM complex GENES & DEVELOPMENT Wen, H., Andrejka, L., Ashton, J., Karess, R., Lipsick, J. S. 2008; 22 (5): 601-614

  Abstract

  The Drosophila Myb oncoprotein, the E2F2 transcriptional repressor, and the RBF and Mip130/LIN-9 tumor suppressor proteins reside in a conserved Myb-MuvB (MMB)/dREAM complex. We now show that Myb is required in vivo for the expression of Polo kinase and components of the spindle assembly checkpoint (SAC). Surprisingly, the highly conserved DNA-binding domain was not essential for assembly of Myb into MMB/dREAM, for transcriptional regulation in vivo, or for rescue of Myb-null mutants to adult viability. E2F2, RBF, and Mip130/LIN-9 acted in opposition to Myb by repressing the expression of Polo and SAC genes in vivo. Remarkably, the absence of both Myb and Mip130, or of both Myb and E2F2, caused variegated expression in which high or low levels of Polo were stably inherited through successive cell divisions in imaginal wing discs. Restoration of Myb resulted in a uniformly high level of Polo expression similar to that seen in wild-type tissue, whereas restoration of Mip130 or E2F2 extinguished Polo expression. Our results demonstrate epigenetic regulation of gene expression by Myb, Mip130/LIN-9, and E2F2-RBF in vivo, and also provide an explanation for the ability of Mip130-null mutants to rescue the lethality of Myb-null mutants in vivo.

  View details for DOI 10.1101/gad.1626308

  View details for Web of Science ID 000253779300005

  View details for PubMedID 18316477

  View details for PubMedCentralID PMC2259030

• p53 arrests growth and induces differentiation of v-Myb-transformed monoblasts DIFFERENTIATION Navratilova, J., Horvath, V., Kozubik, A., Lojek, A., Lipsick, J., Smarda, J. 2007; 75 (7): 592-604

  Abstract

  The p53 protein can control cell cycle progression, programmed cell death, and differentiation of many cell types. Ectopic expression of p53 can resume capability of cell cycle arrest, differentiation, and apoptosis in various leukemic cell lines. In this work, we expressed human p53 protein in v-Myb-transformed chicken monoblasts. We found that even this protein possessing only 53% amino acid homology to its avian counterpart can significantly alter morphology and physiology of these cells causing the G2-phase cell cycle arrest and early monocytic differentiation. Our results document that the species-specific differences of the p53 molecules, promoters/enhancers, and co-factors in avian and human cells do not interfere with differentiation- and cell cycle arrest promoting capabilites of the p53 tumor suppressor even in the presence of functional v-Myb oncoprotein. The p53-induced differentiation and cell cycle arrest of v-Myb-transformed monoblasts are not associated with apoptosis suggesting that the p53-driven pathways controlling apoptosis and differentiation/proliferation are independent.

  View details for DOI 10.1111/j.1432-0436.2006.00158.x

  View details for Web of Science ID 000248966900003

  View details for PubMedID 17309603

• Loss of Drosophila Myb interrupts the progression of chromosome condensation NATURE CELL BIOLOGY Manak, R., Wen, H., Van, T., Andrejka, L., Lipsick, J. S. 2007; 9 (5): 581-U177

  Abstract

  Completion of chromosome condensation is required before segregation during the mitotic cell cycle to ensure the transmission of genetic material with high fidelity in a timely fashion. In many eukaryotes this condensation is regulated by phosphorylation of histone H3 on Ser 10 (H3S10). This phosphorylation normally begins in the late-replicating pericentric heterochromatin and then spreads to the earlier replicating euchromatin. Here, we show that these phases of condensation are genetically separable in that the absence of Drosophila Myb causes cells to arrest with H3S10 phosphorylation of heterochromatin but not euchromatin. In addition, we used mosaic analysis to demonstrate that although the Myb protein can be removed in a single cell cycle, the failure of chromosome condensation occurs only after many cell divisions in the absence of Myb protein. The Myb protein is normally located in euchromatic but not heterochromatic regions of the nucleus, suggesting that Myb may be essential for a modification of euchromatin that is required for the efficient spread of chromosome condensation.

  View details for DOI 10.1038/ncb1580

  View details for Web of Science ID 000246181500017

  View details for PubMedID 17450131

• v-Myb represses the transcription of Ets-2 ONCOGENE Wang, D., Sevcikova, S., Wen, H., Roberts, S., Lipsick, J. S. 2007; 26 (8): 1238-1244

  Abstract

  The v-Myb oncogene causes monoblastic leukemia and transforms only myelomonocytic cells in culture. The v-Myb protein is nuclear and binds to specific DNA sequences. To identify genes regulated by v-Myb, we utilized primary cells transformed by a retrovirus encoding a v-Myb-estrogen receptor (ER) fusion protein. The Ets-2 gene was not expressed in v-Myb-ER transformed cells in the presence of estradiol, but was expressed within 4 h after estradiol withdrawal. The expression of Ets-2 also increased dramatically following phorbol ester-induced differentiation of the v-Myb-transformed BM2 cell line. Conversely, CRYP-alpha, encoding a transmembrane tyrosine phosphatase, was expressed in the presence but not the absence of estradiol in v-Myb-ER transformed cells. CRYP-alpha was downregulated during the phorbol ester-induced differentiation of BM2 cells. Although LIM-3 expression was estradiol-inducible in v-Myb-ER transformed monoblasts, LIM-3 was expressed neither in primary yolk sac cells transformed by unfused v-Myb nor in BM2 cells. We conclude that although v-Myb has been intensively studied as a transcriptional activator, v-Myb can repress biologically relevant genes such as Ets-2, which promotes macrophage differentiation. In addition, we have shown that some genes that are regulated by a v-Myb-ER fusion protein may not be relevant to the biological function of the unfused v-Myb protein.

  View details for DOI 10.1038/sj.onc.1209868

  View details for Web of Science ID 000244406400014

  View details for PubMedID 16909100

• Myb proteins inhibit fibroblast transformation by v-Rel MOLECULAR CANCER Fu, S., Ganter, B., Lipsick, J. S. 2006; 5

  Abstract

  Genes that cause cancer have been divided into two general classes--oncogenes that act in a dominant fashion to transform normal cells into a malignant state, and tumor suppressor genes that act in a dominant fashion to prevent such transformation. In this report, we demonstrate that both the v-myb retroviral oncogene, which causes leukemic transformation of hematopoietic cells, and the c-myb proto-oncogene can also function as inhibitors of fibroblast transformation by the v-rel oncogene. These results imply that the myb genes can function either as oncogenes or as tumor suppressors in different cellular contexts.

  View details for DOI 10.1186/1476-4598-5-54

  View details for Web of Science ID 000241878700002

  View details for PubMedID 17081304

  View details for PubMedCentralID PMC1635057

• Functional evolution of the vertebrate Myb gene family: B-Myb, but neither A-Myb nor C-Myb, complements drosophila Myb in hemocytes GENETICS Davidson, C. J., Tirouvanziam, R., Herzenberg, L. A., Lipsick, J. S. 2005; 169 (1): 215-229

  Abstract

  The duplication of genes and genomes is believed to be a major force in the evolution of eukaryotic organisms. However, different models have been presented about how duplicated genes are preserved from elimination by purifying selection. Preservation of one of the gene copies due to rare mutational events that result in a new gene function (neofunctionalization) necessitates that the other gene copy retain its ancestral function. Alternatively, preservation of both gene copies due to rapid divergence of coding and noncoding regions such that neither retains the complete function of the ancestral gene (subfunctionalization) may result in a requirement for both gene copies for organismal survival. The duplication and divergence of the tandemly arrayed homeotic clusters have been studied in considerable detail and have provided evidence in support of the subfunctionalization model. However, the vast majority of duplicated genes are not clustered tandemly, but instead are dispersed in syntenic regions on different chromosomes, most likely as a result of genome-wide duplications and rearrangements. The Myb oncogene family provides an interesting opportunity to study a dispersed multigene family because invertebrates possess a single Myb gene, whereas all vertebrate genomes examined thus far contain three different Myb genes (A-Myb, B-Myb, and c-Myb). A-Myb and c-Myb appear to have arisen by a second round of gene duplication, which was preceded by the acquisition of a transcriptional activation domain in the ancestral A-Myb/c-Myb gene generated from the initial duplication of an ancestral B-Myb-like gene. B-Myb appears to be essential in all dividing cells, whereas A-Myb and c-Myb display tissue-specific requirements during spermatogenesis and hematopoiesis, respectively. We now report that the absence of Drosophila Myb (Dm-Myb) causes a failure of larval hemocyte proliferation and lymph gland development, while Dm-Myb(-/-) hemocytes from mosaic larvae reveal a phagocytosis defect. In addition, we show that vertebrate B-Myb, but neither vertebrate A-Myb nor c-Myb, can complement these hemocyte proliferation defects in Drosophila. Indeed, vertebrate A-Myb and c-Myb cause lethality in the presence or absence of endogenous Dm-Myb. These results are consistent with a neomorphic origin of an ancestral A-Myb/c-Myb gene from a duplicated B-Myb-like gene. In addition, our results suggest that B-Myb and Dm-Myb share essential conserved functions that are required for cell proliferation. Finally, these experiments demonstrate the utility of genetic complementation in Drosophila to explore the functional evolution of duplicated genes in vertebrates.

  View details for DOI 10.1534/genetics.104.034132

  View details for Web of Science ID 000227039900018

  View details for PubMedID 15489525

  View details for PubMedCentralID PMC1448883

• synMuv verite - Myb comes into focus GENES & DEVELOPMENT Lipsick, J. S. 2004; 18 (23): 2837-2844

  View details for DOI 10.1101/gad.1274804

  View details for Web of Science ID 000225550100001

  View details for PubMedID 15574590

• Fluorescence-activated cell sorting (FACS) of Drosophila hemocytes reveals important functional similarities to mammalian leukocytes PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Tirouvanziam, R., Davidson, C. J., Lipsick, J. S., Herzenberg, L. A. 2004; 101 (9): 2912-2917

  Abstract

  Drosophila is a powerful model for molecular studies of hematopoiesis and innate immunity. However, its use for functional cellular studies remains hampered by the lack of single-cell assays for hemocytes (blood cells). Here we introduce a generic method combining fluorescence-activated cell sorting and nonantibody probes that enables the selective gating of live Drosophila hemocytes from the lymph glands (larval hematopoietic organ) or hemolymph (blood equivalent). Gated live hemocytes are analyzed and sorted at will based on precise quantitation of fluorescence levels originating from metabolic indicators, lectins, reporters (GFP and beta-galactosidase) and antibodies. With this approach, we discriminate and sort plasmatocytes, the major hemocyte subset, from lamellocytes, an activated subset present in gain-of-function mutants of the Janus kinase and Toll pathways. We also illustrate how important, evolutionarily conserved, blood-cell-regulatory molecules, such as calcium and glutathione, can be studied functionally within hemocytes. Finally, we report an in vivo transfer of sorted live hemocytes and their successful reanalysis on retrieval from single hosts. This generic and versatile fluorescence-activated cell sorting approach for hemocyte detection, analysis, and sorting, which is efficient down to one animal, should critically enhance in vivo and ex vivo hemocyte studies in Drosophila and other species, notably mosquitoes.

  View details for DOI 10.1073/pnas.0308734101

  View details for Web of Science ID 000220065300049

  View details for PubMedID 14976247

  View details for PubMedCentralID PMC365719

• Role for a Drosophila Myb-containing protein complex in site-specific DNA replication NATURE Beall, E. L., Manak, J. R., Zhou, S., Bell, M., Lipsick, J. S., Botchan, M. R. 2002; 420 (6917): 833-837

  Abstract

  There is considerable interest in the developmental, temporal and tissue-specific patterns of DNA replication in metazoans. Site-specific DNA replication at the chorion loci in Drosophila follicle cells leads to extensive gene amplification, and the organization of the cis-acting DNA elements that regulate this process may provide a model for how such regulation is achieved. Two elements important for amplification of the third chromosome chorion gene cluster, ACE3 and Ori-beta, are directly bound by Orc (origin recognition complex), and two-dimensional gel analysis has revealed that the primary origin used is Ori-beta (refs 7-9). Here we show that the Drosophila homologue of the Myb (Myeloblastosis) oncoprotein family is tightly associated with four additional proteins, and that the complex binds site-specifically to these regulatory DNA elements. Drosophila Myb is required in trans for gene amplification, showing that a Myb protein is directly involved in DNA replication. A Drosophila Myb binding site, as well as the binding site for another Myb complex member (p120), is necessary in cis for replication of reporter transgenes. Chromatin immunoprecipitation experiments localize both proteins to the chorion loci in vivo. These data provide evidence that specific protein complexes bound to replication enhancer elements work together with the general replication machinery for site-specific origin utilization during replication.

  View details for DOI 10.1038/nature01228

  View details for Web of Science ID 000179897300061

  View details for PubMedID 12490953

• Mutation of the Drosophila homologue of the Myb protooncogene causes genomic instability PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Manak, J. R., Mitiku, N., Lipsick, J. S. 2002; 99 (11): 7438-7443

  Abstract

  Vertebrates have three related Myb genes. The c-Myb protooncogene is required for definitive hematopoiesis in mice and when mutated causes leukemias and lymphomas in birds and mammals. The A-Myb gene is required for spermatogenesis and mammary gland proliferation in mice. The ubiquitously expressed B-Myb gene is essential for early embryonic development in mice and is directly regulated by the p16/cyclin D/Rb family/E2F pathway along with many critical S-phase genes. Drosophila has a single Myb gene most closely related to B-Myb. We have isolated two late-larval lethal alleles of Drosophila Myb. Mutant imaginal discs show an increased number of cells arrested in M phase. Mutant mitotic cells display a variety of abnormalities including spindle defects and increased polyploidy and aneuploidy. Remarkably, some mutant cells have an aberrant S- to M-phase transition in which replicating chromosomes undergo premature histone phosphorylation and chromosomal condensation. These results suggest that the absence of Drosophila Myb causes a defect in S phase that may result in M-phase abnormalities. Consistent with a role for Drosophila Myb during S phase, we detected Dm-Myb protein in S-phase nuclei of wild-type mitotic cells as well as endocycling cells, which lack both an M phase and cyclin B expression. Moreover, we found that the Dm-Myb protein is concentrated in regions of S-phase nuclei that are actively undergoing DNA replication. Together these findings imply that Dm-Myb provides an essential nontranscriptional function during chromosomal replication.

  View details for DOI 10.1073/pnas.122231599

  View details for Web of Science ID 000175908600023

  View details for PubMedID 12032301

  View details for PubMedCentralID PMC124249

• Mutational analysis of the transcriptional activation domains of v-Myb ONCOGENE Wang, D. M., Lipsick, J. S. 2002; 21 (10): 1611-1615

  Abstract

  A minimal transcription activation domain of the v-Myb oncoprotein was initially mapped to a central cluster of charged residues using GAL4-Myb fusion proteins. This region has been proposed to interact directly with the CBP co-activator in animal cells. Regions flanking this central domain of v-Myb are required for transcriptional activation by the native, unfused protein in both mammalian cells and in budding yeast. To identify the critical residues for transcriptional activation, we have now subjected the minimal activation domain and flanking regions including the heptad leucine repeat to random PCR-mediated mutagenesis. We found that the entire region examined can endure extensive substitutions without affecting transcriptional activation by v-Myb in budding yeast. The few mutations that did affect transcriptional activation altered acidic residues within the minimal activation domain or the heptad leucine repeat region, rather than leucine residues. Remarkably, there was a strong concordance between transcriptional activation in animal cells and in budding yeast, even though budding yeast have no known homologue of CBP or related co-activators. In contrast, there was not a strong correlation between transcriptional activation and oncogenic transformation.

  View details for DOI 10.1038/sj/onc/1205236

  View details for Web of Science ID 000173956800017

  View details for PubMedID 11896590

• Functional evolution of the Myb oncogene family 2nd International Workshop on Myb Genes (Myb 2000) Lipsick, J. S., Manak, J., Mitiku, N., Chen, C. K., Fogarty, P., Guthrie, E. ACADEMIC PRESS INC ELSEVIER SCIENCE. 2001: 456–58

  Abstract

  Three Myb-related genes (A-Myb, B-Myb, and c- Myb) have been found in all vertebrates examined thus far including mammals, birds, and amphibians. Two invertebrates, the sea urchin and the fruit fly, have only one Myb-related gene. Our laboratory has used Drosophila as a model system to explore the function of its sole Myb gene. We have also reintroduced the three different vertebrate Myb genes into Drosophila in order to begin to understand how their different functions may have arisen following gene duplication during evolution.

  View details for Web of Science ID 000169361800015

  View details for PubMedID 11259167

• BS69, an adenovirus E1A-associated protein, inhibits the transcriptional activity of c-Myb ONCOGENE Ladendorff, N. E., Wu, S., Lipsick, J. S. 2001; 20 (1): 125-132

  Abstract

  The carboxyl terminus of c-Myb contains a negative regulatory domain that is absent in the v-Myb oncoprotein, but conserved among all the known Myb proteins of animals. This domain inhibits transcriptional activation by c-Myb in animal cells, but not in budding yeast, suggesting that additional protein(s) present in animal cells but not yeast are required for this negative regulatory function. A yeast two-hybrid screen identified BS69, an adenovirus E1A-associated protein, as interacting with the carboxy-terminal region of c-Myb. BS69 contains regions of similarity to the PHD finger, the bromodomain, and the MYND domain, all of which are found in other proteins present in high molecular weight complexes that regulate transcription and/or modify chromatin structure. Further study showed that BS69 inhibited the transcriptional activity of c-Myb, that this inhibition was specific, that it mapped to the carboxyl termini of the two proteins and that it was dose-dependent. A direct interaction between these two proteins was observed in vitro. Furthermore, the 289R E1A protein could inhibit the BS69-mediated decrease in transcriptional activation by c-Myb. By analogy with the inhibition of the Rb/E2F regulatory axis by E1A, we propose that a BS69/Myb regulatory circuit may also be a target of disruption during oncogenesis. Oncogene (2001) 20, 125 - 132.

  View details for Web of Science ID 000166361400014

  View details for PubMedID 11244510

• BS69, an adenovirus E1A-associated protein binds to and inhibits the transcriptional activity of c-Myb Oncogene Lipsick, J.S, Ladendorff, N.E., Wu, S. 2001; 20
• Transcriptional activation by the Myb proteins requires a specific local promoter structure FEBS LETTERS Ganter, B., Chao, S. T., Lipsick, J. S. 1999; 460 (3): 401-410

  Abstract

  The biological effects of the cellular c-Myb and the viral v-Myb proteins are strikingly different. While c-Myb is indispensable for normal hematopoiesis, v-Myb induces acute leukemia. The v-Myb DNA-binding domain (DBD) differs from that of c-Myb mainly by deletion of the first of three repeats which correlates with efficient oncogenic transformation and a decrease in DNA-binding activity. To investigate the difference in DNA-binding and transcriptional activation, oligonucleotide selection and electrophoretic mobility shift assays were employed. The v-Myb DBD (R2R3) shows an intrinsic DNA-binding specificity for an AT-rich downstream extension of the Myb recognition element (MRE) PyAAC(T)/(G)G for efficient binding to this site, whereas R1 within the c-Myb DBD allows for more flexibility for this downstream extension. Therefore, due to the presence of repeat R1, c-Myb can bind to a greater number of target sites. The intrinsic DNA-binding specificity of R2R3 is further supported with the results from in vivo transcriptional activation experiments which demonstrated that both the v-Myb and c-Myb DBDs require an extension of the MRE (motif #1) by a downstream T-stretch (motif #2) for full activity. Surprisingly, the T-stretch improves binding when present on either strand, but is required on a specific strand for transcriptional activation.

  View details for Web of Science ID 000083663500003

  View details for PubMedID 10556506

• Functional analysis of carboxy-terminal deletion mutants of c-Myb JOURNAL OF VIROLOGY Wang, D. M., Dubendorff, J. W., Woo, C. H., Lipsick, J. S. 1999; 73 (7): 5875-5886

  Abstract

  The c-myb gene is implicated in the differentiation and proliferation of hematopoietic cells. Truncations of the N and/or C terminus of c-Myb, found in v-Myb, can potentiate its transforming ability. Two negative regulatory subregions, located in the C terminus, were mapped previously by using GAL4-c-Myb fusion proteins in transient transfection assays for the transcriptional activation of a GAL4-responsive reporter gene. To dissect the C terminus of c-Myb in terms of its involvement in transcriptional activation and oncogenic transformation, a series of C-terminal deletion mutants of c-Myb were analyzed. In addition, linker insertion mutants within the transactivation domain and/or heptad leucine repeat of c-Myb were examined along with those deletion mutants. In this study, we demonstrated that the removal of both of the two previously mapped negative regulatory subregions from the native form of c-Myb not only supertransactivates a Myb-responsive reporter gene but also potentiates its transforming ability in culture. However, in contrast to previous results, cells transformed by all of the mutants analyzed here except v-Myb itself exhibited the same phenotype as those transformed by c-Myb. The proliferating cells were bipotenial and differentiated into both the granulocytic and monocytic lineages. This result implies that the C terminus of c-Myb alone has no effect on the lineage determination. Finally, the transactivation activities of these mutants correlated with their transforming activities when a mim-1 reporter gene was used but not when a model promoter containing five tandem Myb-binding sites was used. In particular, a very weakly transforming mutant with a linker insertion in the heptad leucine repeat superactivated the model promoter but not the mim-1 reporter gene.

  View details for Web of Science ID 000080813500070

  View details for PubMedID 10364339

  View details for PubMedCentralID PMC112648

• Transcriptional regulation by the carboxyl terminus of c-Myb depends upon both the Myb DNA-binding domain and the DNA recognition site ONCOGENE Dubendorff, J. W., Lipsick, J. S. 1999; 18 (23): 3452-3460

  Abstract

  The c-Myb protein binds to DNA, can regulate transcription, and is required for normal hematopoiesis in vertebrates. Either amino- or carboxy-terminal truncation of this protein is required for efficient oncogenic activation. Previous studies have shown that the carboxyl terminus of c-Myb that is deleted in v-Myb contains negative regulatory domains. We now demonstrate that specific mutations within this carboxyl terminus result in greater transcriptional activation than truncation of the entire carboxyl terminus. Furthermore, this increased transcriptional activation depends upon the presence of the highly conserved Myb DNA-binding domain and is also dependent upon the nature of the Myb-binding sites within the target promoter. In a similar fashion, an activating mutation within the heptad leucine repeat region of c-Myb that is also present in v-Myb functions only in conjunction with the Myb DNA-binding domain and with particular Myb-binding sites. These results suggest a model in which multiple domains of the c-Myb protein are highly interdependent for transcriptional regulation. These interactions are promoter-specific and are not well modeled by heterologous fusion proteins.

  View details for Web of Science ID 000080850600003

  View details for PubMedID 10376523

• Transformation by v-Myb ONCOGENE Lipsick, J. S., Wang, D. M. 1999; 18 (19): 3047-3055

  Abstract

  The v-myb oncogene of the avian myeloblastosis virus (AMV) is unique among known oncogenes in that it causes only acute leukemia in animals and transforms only hematopoietic cells in culture. AMV was discovered in the 1930s as a virus that caused a disease in chickens that is similar to acute myelogenous leukemia in humans (Hall et al., 1941). This avian retrovirus played an important role in the history of cancer research for two reasons. First, AMV was used to demonstrate that all oncogenic viruses did not contain a single cancer-causing principle. In particular, although both Rous sarcoma virus (RSV) and AMV could replicate in cultures of either embryonic fibroblasts or hematopoietic cells, RSV could transform only fibroblasts whereas AMV could transform only hematopoietic cells (Baluda, 1963; Durban and Boettiger, 1981a). Second, chickens infected with AMV develop remarkably high white counts and therefore their peripheral blood contains remarkably large quantities of viral particles (Beard, 1963). For this reason AMV was often used as a prototypic retrovirus in order to study viral assembly and later to produce large amounts of reverse transcriptase for both research and commercial purposes. Following the discovery of the v-src oncogene of RSV and the demonstration that it arose from the normal c-src proto-oncogene, a number of acute leukemia viruses were analysed by similar techniques and found to also contain viral oncogenes of cellular origin (Roussel et al., 1979). In the case of AMV, it was shown that almost the entire retroviral env gene had been replaced by a sequence of cellular origin (initially called mab or amv, but later renamed v-myb) (Duesberg et al., 1980; Souza et al., 1980). Remarkably, sequences contained in this myb oncogene were shared between AMV and the avian E26 leukemia virus, but were not contained in any other acutely transforming retroviruses. In addition, the E26 virus contained a second sequence of cellular origin (ets) that was unique. The E26 leukemia virus was first described in the 1960s and causes an acute erythroblastosis in chickens, more reminiscent of the disease caused by avian erythroblastosis virus (AEV) than by AMV (Ivanov et al., 1962).

  View details for Web of Science ID 000080387800014

  View details for PubMedID 10378700

• Myb and oncogenesis ADVANCES IN CANCER RESEARCH, VOL 76 Ganter, B., Lipsick, J. S. 1999; 76: 21-60

  View details for Web of Science ID 000080337300002

  View details for PubMedID 10218098

• Overexpression of an alternatively spliced form of c-Myb results in increases in transactivation and transforms avian myelomonoblasts JOURNAL OF VIROLOGY Woo, C. H., Sopchak, L., Lipsick, J. S. 1998; 72 (8): 6813-6821

  Abstract

  An alternatively spliced form of c-myb exists that encodes an additional 120 amino acids in chicken and 121 amino acids in human and mouse. These amino acids are encoded by an additional exon, termed exon 9A. This exon is not present in v-myb, and proteins containing these amino acids have never been tested for oncogenic transformation. A series of myb constructs was therefore created in order to compare the functions of Myb proteins on the basis of their inclusion or exclusion of the amino acids encoded by exon 9A (E9A). We found that the presence of E9A resulted in a robust increase in transactivation for full-length c-Myb (CCC), as well as the singly truncated derivatives dCC and CCd, while doubly truncated Myb proteins v-Myb (dVd) and dCd did not exhibit any differences in transactivation. The increase in transactivation requires the Myb DNA-binding domain. When the leukemic transformation by the Myb proteins was tested, it was found that cells transformed by dVd resembled monoblasts, while cells transformed by CCC and its derivatives, dCd, dCC, and CCd, resembled myelomonoblasts. Despite differences in the morphology of the hematopoietic cells, the cell surface phenotypes and cell cycle profiles of transformed cells did not change for each pair of Myb proteins in the presence or absence of E9A. Thus, there was no direct correlation between the level of transcriptional activation and the strength of leukemic transformation.

  View details for Web of Science ID 000074730200066

  View details for PubMedID 9658130

  View details for PubMedCentralID PMC109890

• Myb-related Schizosaccharomyces pombe cdc5p is structurally and functionally conserved in eukaryotes MOLECULAR AND CELLULAR BIOLOGY Ohi, R., Feoktistova, A., McCann, S., Valentine, V., Look, A. T., Lipsick, J. S., Gould, K. L. 1998; 18 (7): 4097-4108

  Abstract

  Schizosaccharomyces pombe cdc5p is a Myb-related protein that is essential for G2/M progression. To explore the structural and functional conservation of Cdc5 throughout evolution, we isolated Cdc5-related genes and cDNAs from Saccharomyces cerevisiae, Caenorhabditis elegans, Drosophila melanogaster, and Homo sapiens. Supporting the notion that these Cdc5 gene family members are functionally homologous to S. pombe cdc5(+), human and fly Cdc5 cDNAs are capable of complementing the temperature-sensitive lethality of the S. pombe cdc5-120 mutant. Furthermore, S. cerevisiae CEF1 (S. cerevisiae homolog of cdc5(+)), like S. pombe cdc5(+), is essential during G2/M. The location of the cdc5-120 mutation, as well as mutational analyses of Cef1p, indicate that the Myb repeats of cdc5p and Cef1p are important for their function in vivo. However, we found that unlike in c-Myb, single residue substitutions of glycines for hydrophobic residues within the Myb repeats of Cef1p, which are essential for maintaining structure of the Myb domain, did not impair Cef1p function in vivo. Rather, multiple W-to-G substitutions were required to inactivate Cef1p, and many of the substitution mutants were found to confer temperature sensitivity. Although it is possible that Cef1p acts as a transcriptional activator, we have demonstrated that Cef1p is not involved in transcriptional activation of a class of G2/M-regulated genes typified by SWI5. Collectively, these results suggest that Cdc5 family members participate in a novel pathway to regulate G2/M progression.

  View details for Web of Science ID 000074380100048

  View details for PubMedID 9632794

  View details for PubMedCentralID PMC108994

• D-type cyclins repress transcriptional activation by the v-Myb but not the c-Myb DNA-binding domain EMBO JOURNAL Ganter, B., Fu, S. L., Lipsick, J. S. 1998; 17 (1): 255-268

  Abstract

  The v-Myb DNA-binding domain differs from that of c-Myb mainly by deletion of the first of three repeats. This truncation correlates with efficient oncogenic transformation and a decrease in DNA-binding activity. Here we demonstrate that the D-type cyclins, cyclin D1 and D2 in particular, specifically inhibit transcription when activated through the v-Myb DNA-binding domain, but not the c-Myb DNA-binding domain. Analysis of a cyclin D1 mutant and a dominant-negative CDK4 mutant implied that this repression is independent of complex formation with a CDK partner. Association of cyclin D1 and D2 with the Myb DNA-binding domain could be demonstrated. Increased levels of cyclin D1 and D2 resulted in a stabilization of the Myb proteins, but not in an alteration in binding of the Myb proteins to DNA. These results highlight an unexpected role for cyclin D as a CDK-independent repressor of transcriptional activation by v-Myb but not c-Myb. This differential effect of D-type cyclins on v-Myb and c-Myb might help to explain the mechanism underlying the oncogenic activity of v-Myb, which appears to be a stronger transcriptional activator following the TPA-induced differentiation of transformed monoblasts when cyclin D1 and D2 are down-regulated.

  View details for Web of Science ID 000071708000024

  View details for PubMedID 9427759

  View details for PubMedCentralID PMC1170376

• Myb binding sites within the N-ras promoter repress transcription ONCOGENE Ganter, B., Lipsick, J. S. 1997; 15 (2): 193-202

  Abstract

  In vitro and in vivo methods were combined to determine the function of the three Myb binding sites (NrasI, NrasII and NrasIII) within the promoter region of the mouse N-ras gene. We found that the c-Myb DNA-binding domain can bind with high affinity to NrasI and NrasII, but with a reduced affinity to NrasIII. In contrast, the full length v-Myb protein from BM2 cells only bound to the middle one of the three sites, NrasII. Both c-Myb and v-Myb functioned as repressors and reduce the basal activity of the N-ras promoter by 60%, as determined by transient transfection experiments using different regions of the N-ras promoter. This repression required a functional Myb DNA-binding domain and could not be overcome by fusion to the potent VP16 activation domain. In electrophoretic mobility shift assays, the v-Myb protein is shown to be present in different conformations depending on its binding to the NrasII or the mim-1A site. The v-Myb conformation is thus suggested to play a critical role in the regulation of v-Myb activity.

  View details for Web of Science ID A1997XK37200007

  View details for PubMedID 9244354

• Cells transformed by a v-Myb-estrogen receptor fusion differentiate into multinucleated giant cells JOURNAL OF VIROLOGY Engelke, U., Wang, D. M., Lipsick, J. S. 1997; 71 (5): 3760-3766

  Abstract

  In order to make conditional alleles of the v-myb oncogene, we constructed and tested avian retroviruses which produce a number of different fusion proteins between v-Myb and the human estrogen receptor (ER). We found that the portion of the ER used in making these fusions profoundly affected their transcriptional activation. However, all the fusions tested were only weakly transforming in embryonic yolk sac assays and there was no direct correlation between the level of transcriptional activation and strength of oncogenic transformation. Nevertheless, transformation by a v-Myb-ER fusion was estrogen dependent, and upon withdrawal of the hormone, monocytic-lineage cells differentiated into multinucleated giant cells. Surprisingly, the withdrawal of estrogen caused a dramatic increase in the stability of the fusion protein, although it remained unable to promote cell growth or block differentiation.

  View details for Web of Science ID A1997WT18900047

  View details for PubMedID 9094651

  View details for PubMedCentralID PMC191526

• Constitutive expression of full-length c-Myb transforms avian cells characteristic of both the monocytic and granulocytic lineages CELL GROWTH & DIFFERENTIATION Fu, S. L., Lipsick, J. S. 1997; 8 (1): 35-45

  Abstract

  Both viral Myb (v-Myb) and cellular Myb (c-Myb) are nuclear sequence-specific DNA-binding proteins that can function as transcriptional activators. v-Myb, encoded by avian myeloblastosis virus, induces acute monoblastic leukemia in chickens and transforms avian myelomonocytic cells in culture. The normal c-Myb protein is essential for hematopoietic development. Previous reports suggested that truncation of c-Myb is required for oncogenic transformation of avian myelomonocytic cells in culture. In this study, we demonstrate that constitutive expression of full-length c-Myb can transform avian myelomonocytic cells isolated from embryonic yolk sacs by using a strategy to enhance the efficiency of infection and/or expression of c-myb-containing viruses. c-Myb-transformed myelomonocytic cells display a different phenotype than cells transformed by v-MybAMV or other Myb mutants. c-Myb-transformed yolk sac cells are heterogeneous populations with characteristics of both the macrophage and granulocyte lineages. Our results demonstrate that constitutive expression of full-length c-Myb is sufficient to activate its oncogenic potential, but that the target cells for c-Myb are relatively rare and presumably quite immature.

  View details for Web of Science ID A1997WB77900005

  View details for PubMedID 8993832

• FAETL motif required for leukemic transformation by v-Myb JOURNAL OF VIROLOGY Fu, S. L., Lipsick, J. S. 1996; 70 (8): 5600-5610

  Abstract

  The nuclear protein v-Myb, encoded by the avian myeloblastosis virus (AMV), can induce acute monoblastic leukemia in vivo and transform chicken myelomonocytic cells in culture. The N terminus of v-Myb functions as the DNA-binding domain, and multiple central and C-terminal regions of this protein have been reported to function in transcriptional activation of model reporter genes. We showed previously that a C-terminal domain (amino acids 296 to 371) is required for transcriptional activation and transformation of primary chicken myelomonocytic cells. In this study, we have now analyzed a series of C-terminal mutants of v-Myb to further investigate this domain. A strong correlation was observed between transcriptional activation and leukemic transformation by this series of mutants. Furthermore, deletion analyses demonstrate that the C-terminal 41 amino acids of v=MybAMV (amino acids 331 to 371 of the Myb portion) are nonessential whereas further deletion of amino acids 321 to 330 (EFAETLQLID) results in a nonfunctional protein. Hence, we defined a 10-amino-acid subregion (the "FAETL" motif) required for transcriptional activation and oncogenic transformation by v-Myb Amv. The FAETL region is part of a putative leucine zipper structure and lies near a cluster of phosphorylation sites. Our analysis of mutants with substitutions of the zipper leucines or multiple adjacent phosphorylation sites demonstrates that the function of the FAETL motif is not dependent on an intact leucine zipper structure or adjacent phosphorylation sites. The study of GAL4-Myb fusions suggests that this region is important in maintaining a fully functional conformation of v-Myb. The putative leucine zipper structure has previously been proposed to exert inhibitory effects on c-Myb because its mutation caused increased transcriptional transactivation and transformation. Interestingly, our results show that this region is essential for the functions of v-Myb without requiring a heptad leucine repeat.

  View details for Web of Science ID A1996UX58200085

  View details for PubMedID 8764074

  View details for PubMedCentralID PMC190520

• One billion years of Myb ONCOGENE Lipsick, J. S. 1996; 13 (2): 223-235

  Abstract

  The v-myb oncogene of the avian myeloblastosis virus has led to the discovery of a large and growing family of myb-related genes in a wide variety of eukaryotes including animals, plants, fungi and slime molds. The Myb-related proteins contain a highly conserved sequence, often present in multiple tandem repeats which constitute a DNA-binding domain. These proteins generally function in the regulation of cell growth and differentiation, often by coregulating gene expression along with DNA-binding proteins of other classes. This review focuses on the evolution of the myb gene family and the role of these genes in development.

  View details for Web of Science ID A1996VA25200001

  View details for PubMedID 8710361

• DISSOCIATION OF TRANSCRIPTIONAL ACTIVATION AND ONCOGENIC TRANSFORMATION BY V-MYB ONCOGENE CHEN, R. H., Fields, S., Lipsick, J. S. 1995; 11 (9): 1771-1779

  Abstract

  The nuclear oncoprotein v-Myb is a transcriptional activator in both animal cells and the budding yeast Saccharomyces cerevisiae. Previous studies have suggested that an acidic domain of approximately 50 amino acids (amino acids 204-254 of v-Myb) is necessary and sufficient for transcriptional activation by v-Myb, c-Myb and GAL4-Myb fusion proteins. However, we find that first, none of the acidic residues within this region is essential for transcriptional activation in either animal cells or yeast. Second, transcriptional activation requires cooperation among multiple domains of v-Myb. In animal cells, transcriptional activation by v-Myb requires a central domain (amino acids 234-295), a C-terminal domain (amino acids 295-356), plus either of two more N-terminal domains (amino acids 163-197 or 198-232); in yeast, it requires the central domain plus either the C-terminal domain or a more N-terminal domain (amino acids 163-233). Third, although various subsets of these domains are sufficient for transcriptional activation by v-Myb, all of the domains must be present for transformation of primary hematopoietic cells. These results demonstrate that transcriptional activation by v-Myb is not sufficient for oncogenic transformation.

  View details for Web of Science ID A1995TD09400012

  View details for PubMedID 7478605

• BYPASS OF TPA-INDUCED DIFFERENTIATION AND CELL-CYCLE ARREST BY THE C-MYB DNA-BINDING DOMAIN ONCOGENE Engelke, U., Smarda, J., Lipsick, J. S. 1995; 11 (4): 735-741

  Abstract

  Monoblasts transformed by v-Myb can be induced to differentiate into macrophages by treatment with phorbol ester (TPA). This differentiation occurs during both the G1 and the G2 phases of the cell cycle and is accompanied by cell cycle arrest. The introduction of a protein consisting of the three repeats (3R) of the c-Myb DNA-binding domain permits the by-pass of this phorbol ester-induced differentiation and cell cycle arrest. In particular, monoblasts which express the 3R protein progress through both the G1/S and G2/M transitions in the presence of phorbol ester. However, the 3R protein contains no detectable transcriptional activation domain. These results demonstrate that the c-Myb DNA-binding domain can regulate the cell cycle without functioning as a direct transcriptional activator.

  View details for Web of Science ID A1995RQ46900015

  View details for PubMedID 7651737

• RETINOIC ACID RECEPTOR-ALPHA SUPPRESSES TRANSFORMATION BY V-MYB MOLECULAR AND CELLULAR BIOLOGY Smarda, J., Sugarman, J., Glass, C., Lipsick, J. 1995; 15 (5): 2474-2481

  Abstract

  Retinoic acid (RA) is capable of inducing the differentiation of various myelomonocytic cell lines. During this differentiation process, the levels of c-myb expression decline, suggesting that the RA receptor (RAR) may act in part by down-regulating this proto-oncogene. We have now investigated whether the RAR can also inhibit the function of Myb proteins themselves. We have found that transcriptional activation of a Myb-responsive reporter gene can be inhibited by RA in a human monocytic cell line. This inhibition could not be overcome by the expression of exogenous Myb. The RAR did not interfere with DNA binding by Myb proteins in vitro, suggesting that the functional inhibition occurs at the level of transcriptional activation. To determine the biological relevance of the inhibition of Myb proteins by the RAR, we have used v-myb-transformed monoblasts. These cells differentiate into macrophages in the presence of phorbol ester (tetradecanoyl phorbol acetate [TPA]) but are normally unresponsive to RA treatment. The introduction of an inducible, exogenous RAR alpha into v-myb-transformed monoblasts permitted an RA-dependent differentiation into macrophage-like cells similar to those induced by TPA. These results demonstrate that transformation by v-myb is recessive to RAR alpha and imply that many types of non-RA-responsive leukemia cells may become responsive following the introduction of the RAR.

  View details for Web of Science ID A1995QU66500017

  View details for PubMedID 7739532

  View details for PubMedCentralID PMC230477

• WEAK TRANSCRIPTIONAL ACTIVATION IS SUFFICIENT FOR TRANSFORMATION BY V-MYB VIROLOGY Engelke, U., Whittaker, L., Lipsick, J. S. 1995; 208 (2): 467-477

  Abstract

  The v-myb oncogene causes monoblastic leukemia in chickens and transforms avian myelomonocytic cells in vitro, v-Myb is a short-lived nuclear protein which binds to DNA in a sequence-specific manner and can activate gene expression in transient DNA transfections. Analysis of a series of v-Myb mutants has shown that the ability to activate transcription appears to be required for leukemic transformation. We have systematically investigated transcriptional activation by v-Myb and have made several new observations: (i) v-Myb is a very weak activator when compared to GAL4; (ii) very weak transcriptional activation by v-Myb is sufficient for transformation, whereas very strong transcriptional activation by a v-Myb-VP16 fusion protein is not; and (iii) v-Myb can activate transcription by two genetically distinct mechanisms, only one of which requires the presence of Myb-binding sites.

  View details for Web of Science ID A1995QZ40000007

  View details for PubMedID 7747419

• MUTATIONS IN THE DNA-BINDING AND TRANSCRIPTIONAL ACTIVATION DOMAINS OF V-MYB COOPERATE IN TRANSFORMATION JOURNAL OF VIROLOGY DINI, P. W., ELTMAN, J. T., Lipsick, J. S. 1995; 69 (4): 2515-2524

  Abstract

  The v-Myb protein encoded by avian myeloblastosis virus causes oncogenic transformation of monoblastic cells committed to the monocyte/macrophage lineage. v-Myb is a doubly truncated form of its normal cellular counterpart, c-Myb. In addition to its N- and C-terminal deletions, v-Myb contains a number of amino acid substitutions relative to c-Myb. We have previously shown that neither overexpression of c-Myb nor introduction of these amino acid substitutions into c-Myb is sufficient for transformation of myelomonocytic cells. However, a doubly truncated form of c-Myb which lacked these substitutions transformed myeloblastic cells that appeared to be committed to the granulocytic pathway. We demonstrate here that mutations in both the DNA-binding and transcriptional activation domains of v-Myb are required for transformation of rapidly growing monoblasts rather than more slowly growing myeloblasts. These rapidly growing monoblasts do not express mim-1, a target gene for the Gag-Myb-Ets protein of E26 leukemia virus, or C/EBP proteins which cooperate with Myb to activate mim-1 expression. Furthermore, v-Myb proteins which contain both sets of these mutations are weaker transcriptional activators relative to proteins which lack these mutations. These results support a model in which amino acid substitutions in v-Myb have been selected for their ability to activate only a subset of those genes which can be activated by a doubly truncated form of c-Myb. In particular, mim-1 appears to represent a class of genes whose expression was selected against during the development of an increasingly virulent strain of avian myeloblastosis virus by passage in animals.

  View details for Web of Science ID A1995QL68300066

  View details for PubMedID 7884901

  View details for PubMedCentralID PMC188928

• TRANSFORMATION OF MYELOMONOCYTIC CELLS BY THE AVIAN-MYELOBLASTOSIS VIRUS IS DETERMINED BY THE V-MYB ONCOGENE, NOT BY THE UNIQUE LONG TERMINAL REPEATS OF THE VIRUS JOURNAL OF VIROLOGY Engelke, U., Lipsick, J. S. 1994; 68 (4): 2752-2755

  Abstract

  The avian myeloblastosis virus (AMV) induces acute monoblastic leukemia in chickens and transforms only myelomonocytic cells in vitro. The long terminal repeat (LTR) regulatory region of AMV is unique among the known classes of avian retrovirus LTRs. We demonstrate that the substitution of the AMV LTRs by Rous sarcoma virus LTRs did not alter the cell type specificity or the transforming ability of the virus.

  View details for Web of Science ID A1994NA30700079

  View details for PubMedID 8139052

  View details for PubMedCentralID PMC236755

• C-MYB PREVENTS TPA-INDUCED DIFFERENTIATION AND CELL-DEATH IN V-MYB TRANSFORMED MONOBLASTS ONCOGENE Smarda, J., Lipsick, J. S. 1994; 9 (1): 237-245

  Abstract

  Myelomonocytic cells transformed by v-Myb or altered forms of c-Myb do not contain the full-length c-Myb protein found in most immature hematopoietic cells. To determine if c-Myb was a dominant inhibitor of v-Myb, we have induced the synthesis of full-length c-Myb in monoblasts transformed by v-Myb. We found that although some morphological changes occurred, the presence of both c-Myb and v-Myb was compatible with cell growth. However, the response to phorbol ester (TPA) was significantly altered by c-Myb. Monoblasts transformed by v-Myb can be induced to differentiate into macrophages by treatment with TPA. This process is accompanied by a significant amount of cell death. However, when c-Myb was made TPA-inducible in these cells, TPA-induced differentiation into macrophages was blocked and cell death was prevented. These results demonstrate a significant difference in the biological effects of v-Myb and c-Myb in transformed myelomonocytic cells.

  View details for Web of Science ID A1994MW24700029

  View details for PubMedID 8302585

• DICISTRONIC SELECTION FOR NUCLEAR PROTEINS IN LIVING ANIMAL-CELLS GENE Smarda, J., Lipsick, J. S. 1993; 137 (1): 145-149

  Abstract

  We present an expression/selection system designed for the purification of cell lines inducibly expressing genes coding for unselectable proteins by using dicistronic selection for the cell surface marker CD4. This system enabled us to establish and purify c-myb expressing variants of the v-myb transformed chicken monoblast cell line BM2 with high efficiency.

  View details for Web of Science ID A1993MW21700021

  View details for PubMedID 8282197

• INDIVIDUAL REPEATS OF DROSOPHILA MYB CAN FUNCTION IN TRANSFORMATION BY V-MYB JOURNAL OF VIROLOGY Bin, X., Lipsick, J. S. 1993; 67 (12): 7332-7339

  Abstract

  The v-Myb protein binds to specific DNA sequences and can regulate gene expression. The DNA-binding domain of v-Myb contains the second and third of the three highly conserved tandem repeats found in c-Myb. In general, the ability of mutant forms of v-Myb to transform correlates with their ability to trans activate transcription. Two mutations within the DNA-binding domain of v-Myb which preserve DNA binding in vitro but fail to trans activate or transform have been described. These results suggested that this highly conserved domain might function in specific protein-protein interactions, as well as in DNA binding. We therefore tested the ability of a related protein domain from Drosophila melanogaster to substitute functionally for the homologous region of v-Myb. We found that either the second or third repeat of Drosophila Myb, but not both, could function in trans-activation and transformation by v-Myb. The hybrid containing both the second and third repeats of Drosophila Myb bound to DNA but failed to trans activate transcription either in the context of v-Myb or as a v-Myb-VP16 fusion protein. These results demonstrate that although the protein-DNA contacts made by the Myb repeats have been conserved during the evolution of animals, the protein-protein interactions have diverged.

  View details for Web of Science ID A1993MG30700050

  View details for PubMedID 8230457

  View details for PubMedCentralID PMC238197

• ONCOGENIC TRUNCATION OF THE 1ST REPEAT OF C-MYB DECREASES DNA-BINDING IN-VITRO AND IN-VIVO MOLECULAR AND CELLULAR BIOLOGY DINI, P. W., Lipsick, J. S. 1993; 13 (12): 7334-7348

  Abstract

  Oncogenic activation of c-Myb in both avian and murine systems often involves N-terminal truncation. In particular, the first of three DNA-binding repeats in c-Myb has been largely deleted during the genesis of the v-myb oncogenes of avian myeloblastosis virus and E26 avian leukemia virus. This finding suggests that the first DNA-binding repeat may have an important role in cell growth control. We demonstrate that truncation of the first DNA-binding repeat of c-Myb is sufficient for myeloid transformation in culture, but deletion of the N-terminal phosphorylation site and adjacent acidic region is not. Truncation of the first repeat decreases the ability of a Myb-VP16 fusion protein to trans activate the promoter of a Myb-inducible gene (mim-1) involved in differentiation. Moreover, truncation of the first repeat decreases the ability of the Myb protein to bind DNA both in vivo and in vitro. These results suggest that N-terminal mutants of c-Myb may transform by regulating only a subset of those genes normally regulated by c-Myb.

  View details for Web of Science ID A1993MJ34100015

  View details for PubMedID 8246954

  View details for PubMedCentralID PMC364804

• C-MYB AND V-MYB ARE DIFFERENTIALLY PHOSPHORYLATED BY P42MAPK IN-VITRO ONCOGENE Aziz, N., Wu, J., Dubendorff, J. W., Lipsick, J. S., Sturgill, T. W., Bender, T. P. 1993; 8 (8): 2259-2265

  Abstract

  The product of the c-myb proto-oncogene is a highly conserved transcription factor that has been shown to function as both a transactivator and repressor. The v-myb oncogenes of E26 leukemia virus and avian myeloblastosis virus (AMV) encode proteins truncated at both the amino and carboxy termini, deleting portions of the DNA-binding and negative regulatory domains present in c-Myb. Similar truncations of c-Myb alter its function, suggesting that the viral proteins lack important regulatory sequences. Interestingly, eight potential sites of phosphorylation by proline-directed protein kinases conserved between the avian, murine and human Myb proteins are clustered in or near the negative regulatory domain of c-Myb. The majority of these sites are deleted in both the E26 and AMV viral proteins. In this paper we show that one proline-directed protein kinase, p42mapk, phosphorylates bacterially synthesized avian and murine c-Myb but not AMV v-Myb in vitro. We find that p42mapk phosphorylates c-Myb on serine and threonine, but not on tyrosine. Furthermore, deletion analysis indicates that the sites of phosphorylation map to the C-terminal negative regulatory domain. We speculate that the inability of v-Myb to be phosphorylated by p42mapk may contribute to its oncogenic properties.

  View details for Web of Science ID A1993LP17100028

  View details for PubMedID 8336948

• DIFFERENTIAL TRANSCRIPTIONAL ACTIVATION BY V-MYB AND C-MYB IN ANIMAL-CELLS AND SACCHAROMYCES-CEREVISIAE MOLECULAR AND CELLULAR BIOLOGY CHEN, R. H., Lipsick, J. S. 1993; 13 (7): 4423-4431

  Abstract

  The v-myb oncogene and its cellular homolog c-myb encode sequence-specific DNA-binding proteins which regulate transcription from promoters containing Myb-binding sites in animal cells. We have developed a Saccharomyces cerevisiae system to assay transcriptional activation by v-Myb and c-Myb. In yeast strains containing integrated reporter genes, activation was strictly dependent upon both the Myb DNA-binding domain and the Myb recognition element. BAS1, an endogenous Myb-related yeast protein, was not required for transactivation by animal Myb proteins and by itself had no detectable effect on a Myb reporter gene. Deletion analyses demonstrated that a domain of v-Myb C terminal to the previously mapped Myb transcriptional activation domain was required for transactivation in animal cells but not in S. cerevisiae. The same domain is also required for the efficient transformation of myeloid cells by v-Myb. In contrast to results in animal cells, in S. cerevisiae the full-length c-Myb was a much stronger transactivator than a protein bearing the oncogenic N- and C-terminal truncations of v-Myb. These results imply that negative regulation of c-Myb by its own termini requires an additional animal cell protein or small molecule that is not present in S. cerevisiae.

  View details for Web of Science ID A1993LJ36200061

  View details for PubMedID 8321242

• CARBOXY-TERMINAL ELEMENTS OF C-MYB NEGATIVELY REGULATE TRANSCRIPTIONAL ACTIVATION IN CIS AND IN TRANS GENES & DEVELOPMENT Dubendorff, J. W., Whittaker, L. J., ELTMAN, J. T., Lipsick, J. S. 1992; 6 (12B): 2524-2535

  Abstract

  The c-Myb protein plays a key role in normal hematopoiesis, and truncation results in its activation to a transforming protein. Truncation of the c-Myb carboxyl terminus also greatly increases its transcriptional activating activity. The role of specific carboxy-terminal domains in negative regulation was investigated using Myb and Myb fusions with GAL4, LexA, or VP16. Negative regulatory activity of the carboxyl terminus in cis resides in at least two regions. A sequence in one of these regions can also inhibit transcriptional activation by Myb, Myb-VP16, or LexA-Myb proteins in trans. Regulation in trans, or suppression, is independent of c-Myb DNA binding and, therefore, likely involves protein-protein interaction. Suppression does not require the presence of a predicted heptad leucine repeat structure on either molecule. The target of suppression is a sequence that contains part of the minimal Myb transcriptional activation domain. This sequence can confer suppressibility on fusion proteins containing heterologous DNA-binding or transcriptional activation domains.

  View details for Web of Science ID A1992KF41000008

  View details for PubMedID 1340467

• PROTEIN TRUNCATION IS NOT REQUIRED FOR C-MYB PROTOONCOGENE ACTIVITY IN NEURORETINA CELLS JOURNAL OF VIROLOGY GARRIDO, C., Grasser, F., Lipsick, J. S., Stehelin, D., Saule, S. 1992; 66 (11): 6773-6776

  Abstract

  The v-myb oncogene of avian myeloblastosis virus (AMV) differs from its normal cellular counterpart by a truncation at both its amino and carboxyl termini and by a substitution of 11 amino acid residues. We had previously shown that v-myb-containing AMV, in the presence of basic fibroblast growth factor, transformed chicken neuroretina (CNR) cells. To understand the mechanism of c-myb activation, we have tested whether avian retroviruses that express the full-length c-Myb are also active on CNR cells. We have found that c-Myb, like v-Myb, strongly increases the basic fibroblast growth factor response of CNR cells and that these c-myb-expressing cells are able to grow in soft agar in the presence of the growth factor. We have also found that, in contrast to normal or v-myb-expressing AMV-transformed CNR cells, c-Myb-transformed cells express mim-1, a granulocyte-specific gene. However, normal v-Myb- and c-Myb-expressing CNR cells all express the pax-QNR gene, a newly described paired and homeobox-containing gene specifically expressed in the neuroretina. We conclude that, in contrast to what has been described for hematopoietic cells, overexpression of c-Myb is sufficient to activate gene expression and to induce an abnormal behavior of CNR cells.

  View details for Web of Science ID A1992JT70000061

  View details for PubMedID 1404616

• A HIGHLY CONSERVED CYSTEINE IN THE V-MYB DNA-BINDING DOMAIN IS ESSENTIAL FOR TRANSFORMATION AND TRANSCRIPTIONAL TRANSACTIVATION ONCOGENE Grasser, F. A., LaMontagne, K., Whittaker, L., Stohr, S., Lipsick, J. S. 1992; 7 (5): 1005-1009

  Abstract

  The v-Myb protein is nuclear, binds to DNA in a sequence-specific fashion, regulates the transcription of various reporter gene and transforms myelomonocytic cells. Cysteine is one of the most conserved residues during protein evolution and has been implicated in DNA binding, protein-protein interaction and redox regulation of various proteins. Therefore, we have now individually substituted each of the seven cysteines of v-Myb with a serine. All seven mutant proteins bound to DNA when they were expressed in E. coli. However, mutant C65S neither trans-activated transcription in vivo nor transformed myeloid cells, although it was transported into the nucleus. This cysteine is conserved in the Myb-related proteins of animals, plants, yeast and the cellular slime mold Dictyostelium discoideum. The C65S mutation and a nearby codon insertion mutation also abolished trans-activation by fusion proteins containing the v-Myb DNA-binding domain and the strong constitutive activation domain of herpes simplex virus (HSV) VP16. Because this domain of VP16 appears to activate transcription whenever it is bound upstream of an appropriate promoter, these results imply that C65 may be required for high-affinity DNA binding in vivo. In support of this hypothesis, we have also shown that, in contrast to wild-type v-Myb, mutant C65S is unable to block transcription from a reporter gene in which Myb binding sites overlap the initiation site.

  View details for Web of Science ID A1992HP64200022

  View details for PubMedID 1570148

• EXPRESSION OF THE CD4 GENE REQUIRES A MYB TRANSCRIPTION FACTOR MOLECULAR AND CELLULAR BIOLOGY SIU, G., Wurster, A. L., Lipsick, J. S., HEDRICK, S. M. 1992; 12 (4): 1592-1604

  Abstract

  We have analyzed the control of developmental expression of the CD4 gene, which encodes an important recognition molecule and differentiation antigen on T cells. We have determined that the CD4 promoter alone functions at high levels in the CD4+ CD8- mature T cell but not at the early CD4+ CD8+ stage of T-cell development. In addition, the CD4 promoter functions only in T lymphocytes; thus, the stage and tissue specificity of the CD4 gene is mediated in part by its promoter. We have determined that a Myb transcription factor binds to the CD4 promoter and is critical for full promoter function. Thus, Myb plays an important role in the expression of T-cell-specific developmentally regulated genes.

  View details for Web of Science ID A1992HK75100022

  View details for PubMedID 1347906

• THE MYB DNA-BINDING DOMAIN IS HIGHLY CONSERVED IN DICTYOSTELIUM-DISCOIDEUM ONCOGENE STOBERGRASSER, U., BRYDOLF, B., Bin, X., Grasser, F., Firtel, R. A., Lipsick, J. S. 1992; 7 (3): 589-596

  Abstract

  The c-myb proto-oncogene encodes a protein that is highly conserved among birds and mammals. The amino-terminal domain of c-Myb contains three imperfect tandem repeats of approximately 50 amino acids each. This domain is required for DNA binding and has also been conserved to varying degrees in invertebrates, plants and yeast. Given that myb-related genes appear to control cellular differentiation in a variety of eucaryotic systems, the presence of a myb gene in the cellular slime mold Dictyostelium discoideum might provide a tractable system for studying the role of myb in differentiation. Degenerate oligonucleotide primers encoding regions that are highly conserved in the vertebrate and Drosophila Myb DNA-binding domains were used to amplify a related domain from Dictyostelium genomic DNA, which was then used to isolate a genomic clone. The putative DNA-binding domain of Dictyostelium Myb is as closely related to vertebrate c-Myb as is Drosophila Myb (65% identity), whereas the known Myb-related proteins of plants and yeast are more distantly related. The conserved domain of Dictyostelium Myb is capable of binding to the same DNA sequence as the vertebrate and Drosophila Myb proteins. The remainder of the deduced amino acid sequence of Dictyostelium Myb shows no homology to the divergent domains of the known animal, plant and yeast Myb-related proteins. Evolutionary analysis implies that the duplications that generated the repeats of the Myb DNA-binding domain began prior to the divergence of animals, plants, cellular slime molds and yeast.

  View details for Web of Science ID A1992HK00500029

  View details for PubMedID 1549373

• DEFINITION OF FUNCTIONAL DOMAINS IN P135GAG-MYB-ETS AND P48V-MYB PROTEINS REQUIRED TO MAINTAIN THE RESPONSE OF NEURORETINA CELLS TO BASIC FIBROBLAST GROWTH-FACTOR JOURNAL OF VIROLOGY GARRIDO, C., Leprince, D., Lipsick, J. S., Stehelin, D., GOSPODAROWICZ, D., Saule, S. 1992; 66 (1): 160-166

  Abstract

  The v-myb- and v-ets-containing E26 retrovirus induces the proliferation of chicken neuroretina (CNR) cells in minimal medium. Proliferation of E26 CNR cells is strongly stimulated by basic fibroblast growth factor (bFGF). The v-myb-containing avian myeloblastosis virus also induces the proliferation of infected CNR cells stimulated by bFGF. Both E26 CNR and avian myeloblastosis virus CNR cells are able to form colonies in soft agar in the presence of bFGF. This suggests that the v-myb product, a nuclear sequence-specific DNA-binding protein which activates gene expression in transient transfection assays, plays a role in the proliferative response of the infected CNR cells. To determine the structure-function relationships of P135gag-myb-ets and p48v-myb, we have used deletion mutants expressed in retroviral vectors and have analyzed their effect on CNR cell proliferation as well as their effect on the CNR cell response to bFGF. We show that v-ets is not required for bFGF stimulation but increases the proliferation of CNR cells in minimal medium. In the v-myb mutants, the gag sequences derived from the helper virus increase the potency of the myb gene. The carboxyl-terminal domain required for the growth and transformation of myeloid cells and needed for maximal trans-activation in transient DNA transfection assays in fibroblasts was not required for the growth and bFGF response of CNR cells. In contrast, the domain encompassing amino acids 240 to 301 (containing part of the transcriptional activation domain of v-myb) was absolutely required for the response of CNR cells to bFGF and could be functionally replaced by the carboxyl-terminal transcriptional activation domain of the VP16 protein of herpes simplex virus.

  View details for Web of Science ID A1992GU97100020

  View details for PubMedID 1727478

• PROTEIN TRUNCATION IS REQUIRED FOR THE ACTIVATION OF THE C-MYB PROTOONCOGENE MOLECULAR AND CELLULAR BIOLOGY Grasser, F. A., Graf, T., Lipsick, J. S. 1991; 11 (8): 3987-3996

  Abstract

  The protein product of the v-myb oncogene of avian myeloblastosis virus, v-Myb, differs from its normal cellular counterpart, c-Myb, by (i) expression under the control of a strong viral long terminal repeat, (ii) truncation of both its amino and carboxyl termini, (iii) replacement of these termini by virally encoded residues, and (iv) substitution of 11 amino acid residues. We had previously shown that neither the virally encoded termini nor the amino acid substitutions are required for transformation by v-Myb. We have now constructed avian retroviruses that express full-length or singly truncated forms of c-Myb and have tested them for the transformation of chicken bone marrow cells. We conclude that truncation of either the amino or carboxyl terminus of c-Myb is sufficient for transformation. In contrast, the overexpression of full-length c-Myb does not result in transformation. We have also shown that the amino acid substitutions of v-Myb by themselves are not sufficient for the activation of c-Myb. Rather, the presence of either the normal amino or carboxyl terminus of c-Myb can suppress transformation when fused to v-Myb. Cells transformed by c-Myb proteins truncated at either their amino or carboxyl terminus appear to be granulated promyelocytes that express the Mim-1 protein. Cells transformed by a doubly truncated c-Myb protein are not granulated but do express the Mim-1 protein, in contrast to monoblasts transformed by v-Myb that neither contain granules nor express Mim-1. These results suggest that various alterations of c-Myb itself may determine the lineage of differentiating hematopoietic cells.

  View details for Web of Science ID A1991FX72300019

  View details for PubMedID 2072904

• DETERMINANTS OF SEQUENCE-SPECIFIC DNA-BINDING BY P48V-MYB ONCOGENE Garcia, A., LaMontagne, K., REAVIS, D., STOBERGRASSER, U., Lipsick, J. S. 1991; 6 (2): 265-273

  Abstract

  The v-myb oncogene of the avian myeloblastosis virus encodes a nuclear protein, p48v-myb, which binds to DNA in a sequence-specific manner. We have used wild type and mutant forms of this protein expressed in E. coli to study the protein and DNA determinants for sequence-specific DNA-binding. We have shown that only the highly conserved domain at the amino terminus of p48v-myb is required for sequence-specific DNA-binding. However, neither of the tandem 50 amino acid repeats present in this domain is alone sufficient for such binding. We have also demonstrated that p48v-myb can recognize a single consensus myb binding site and appears to interact with DNA as a protein monomer. In addition, we have shown that sequence-specific binding by p48v-myb requires nucleotides which flank the previously reported PyAACT/GG consensus.

  View details for Web of Science ID A1991FZ13400012

  View details for PubMedID 2000220

• TRANSFORMATION BY V-MYB CORRELATES WITH TRANSACTIVATION OF GENE-EXPRESSION MOLECULAR AND CELLULAR BIOLOGY Lane, T., IBANEZ, C., Garcia, A., Graf, T., Lipsick, J. 1990; 10 (6): 2591-2598

  Abstract

  The v-myb oncogene of avian myeloblastosis virus causes acute myelomonocytic leukemia in chickens and transforms avian myeloid cells in vitro. Its protein product p48v-myb is a nuclear, sequence-specific, DNA-binding protein which activates gene expression in transient DNA transfection studies. To investigate the relationship between transformation and trans-activation by v-myb, we constructed 15 in-frame linker insertion mutants. The 12 mutants which transformed myeloid cells also trans-activated gene expression, whereas the 3 mutants which did not transform also did not trans-activate. This implies that trans-activation is required for transformation by v-myb. One of the transformation-defective mutants localized to the cell nucleus but failed to bind DNA. The other two transformation-defective mutants localized to the cell nucleus and bound DNA but nevertheless failed to trans-activate. These latter mutants define two distinct domains of p48v-myb which control trans-activation by DNA-bound protein, one within the amino-terminal DNA-binding domain itself and one in a carboxyl-terminal domain which is not required for DNA binding.

  View details for Web of Science ID A1990DF42500018

  View details for PubMedID 2160580

• TRANS ACTIVATION OF GENE-EXPRESSION BY V-MYB MOLECULAR AND CELLULAR BIOLOGY Ibanez, C. E., Lipsick, J. S. 1990; 10 (5): 2285-2293

  Abstract

  The v-myb oncogene causes acute myelomonocytic leukemia in chickens and transforms avian myeloid cells in vitro. Its product, p48v-myb, is a short-lived nuclear protein which binds DNA. We demonstrate that p48v-myb can function as a trans activator of gene expression in transient DNA transfection assays. trans activation requires the highly conserved amino-terminal DNA-binding domain and the less highly conserved carboxyl-terminal domain of p48v-myb, both of which are required for transformation. Multiple copies of a consensus sequence for DNA binding by p48v-myb inserted upstream of a herpes simplex virus thymidine kinase promoter are strongly stimulatory for transcriptional activation by a v-myb-VP16 fusion protein but not by p48v-myb itself, suggesting that the binding of p48v-myb to DNA may not be sufficient for trans activation.

  View details for Web of Science ID A1990DA19500047

  View details for PubMedID 2325652

• DNA-BINDING ACTIVITY ASSOCIATED WITH THE V-MYB ONCOGENE PRODUCT IS NOT SUFFICIENT FOR TRANSFORMATION JOURNAL OF VIROLOGY Ibanez, C. E., Garcia, A., STOBERGRASSER, U., Lipsick, J. S. 1988; 62 (11): 4398-4402

  Abstract

  The product of the v-myb oncogene of avian myeloblastosis virus is a nuclear protein with an associated DNA-binding activity. We demonstrated that the highly conserved amino-terminal domain of p48v-myb is required for its associated DNA-binding activity. This activity is not required for the nuclear localization of p48v-myb. Furthermore, the associated DNA-binding activity and nuclear localization of p48v-myb together are not sufficient for transformation.

  View details for Web of Science ID A1988Q489900066

  View details for PubMedID 2845150

• STRUCTURAL AND FUNCTIONAL DOMAINS OF THE MYB ONCOGENE - REQUIREMENTS FOR NUCLEAR TRANSPORT, MYELOID TRANSFORMATION, AND COLONY FORMATION JOURNAL OF VIROLOGY Ibanez, C. E., Lipsick, J. S. 1988; 62 (6): 1981-1988

  Abstract

  The v-myb oncogene of avian myeloblastosis virus causes acute myelomonocytic leukemia in vivo and transforms only myeloid cells in vitro. Its product, p48v-myb, is a nuclear protein of unknown function. To determine structure-function relationships for this protein, we constructed a series of deletion mutants of v-myb, expressed them in retroviral vectors, and studied their biochemical and biological properties. We used these mutants to identify two separate domains of p48v-myb which had distinct roles in its accumulation in the cell nucleus. We showed that the viral sequences which normally encode both termini of p48v-myb were dispensible for transformation. In contrast, both copies of the highly conserved v-myb amino-terminal repeat were required for transformation. We also identified a carboxyl-terminal domain of p48v-myb which was required for the growth of v-myb-transformed myeloblasts in soft agar but not for morphological transformation.

  View details for Web of Science ID A1988N407000018

  View details for PubMedID 2835503

• SPECIFIC AMINO-ACID SUBSTITUTIONS ARE NOT REQUIRED FOR TRANSFORMATION BY V-MYB OF AVIAN-MYELOBLASTOSIS VIRUS JOURNAL OF VIROLOGY STOBERGRASSER, U., Lipsick, J. S. 1988; 62 (3): 1093-1096

  Abstract

  The protein product of the v-myb oncogene of avian myeloblastosis virus, p48v-myb, differs structurally in several ways from its normal cellular homolog, p75c-myb. We demonstrated that the 11 specific amino acid substitutions found in two independent molecular clones of this virus were not required for the transformation of myeloblasts by v-myb.

  View details for Web of Science ID A1988M029700058

  View details for PubMedID 2828660

• V-MYB DOES NOT PREVENT THE EXPRESSION OF C-MYB IN AVIAN ERYTHROBLASTS JOURNAL OF VIROLOGY Lipsick, J. S. 1987; 61 (10): 3284-3287

  Abstract

  The v-myb oncogene of avian myeloblastosis virus transforms myeloid cells exclusively, both in vivo and in vitro. The c-myb proto-oncogene from which v-myb arose is expressed at relatively high levels in immature hematopoietic cells of the lymphoid, erythroid, and myeloid lineages but not in myeloblasts transformed by v-myb. This finding suggested that the nuclear v-myb gene product p48v-myb might act directly to inhibit the normal expression of the c-myb gene. I have therefore used a selectable avian retroviral vector to express p48v-myb in avian erythroblasts which normally express high levels of the c-myb gene product p75c-myb. The results demonstrate that p48v-myb and p75c-myb can be coexpressed in the nuclei of cloned cells. Therefore, p48v-myb does not invariably prevent the expression of p75c-myb.

  View details for Web of Science ID A1987J994200044

  View details for PubMedID 3041047

• C-MYB PROTEIN EXPRESSION IS A LATE EVENT DURING LYMPHOCYTE-T ACTIVATION MOLECULAR AND CELLULAR BIOLOGY Lipsick, J. S., Boyle, W. J. 1987; 7 (9): 3358-3360

  Abstract

  The expression of p80c-myb was examined during the activation of resting human T lymphocytes. Before activation, no detectable p80c-myb was present. Synthesis of p80c-myb was observed only after initiation of the S phase of the cell cycle.

  View details for Web of Science ID A1987J716400047

  View details for PubMedID 3118198

• ENV-ENCODED RESIDUES ARE NOT REQUIRED FOR TRANSFORMATION BY P48V-MYB JOURNAL OF VIROLOGY Lipsick, J. S., Ibanez, C. E. 1987; 61 (3): 933-936

  Abstract

  The v-myb oncogene of avian myeloblastosis virus induces acute myeloblastic leukemia in chickens and transforms avian myeloid cells in vitro. The protein product of this oncogene, p48v-myb, is partially encoded by the retroviral gag and env genes. We demonstrated that the env-encoded carboxyl terminus of p48v-myb is not required for transformation. Our results showed, in addition, that a coding region of c-myb which is not essential for transformation was transduced by avian myeloblastosis virus.

  View details for Web of Science ID A1987G030600042

  View details for PubMedID 3027417

• EXPRESSION OF MOLECULAR CLONES OF V-MYB IN AVIAN AND MAMMALIAN-CELLS INDEPENDENTLY OF TRANSFORMATION JOURNAL OF VIROLOGY Lipsick, J. S., Ibanez, C. E., Baluda, M. A. 1986; 59 (2): 267-275

  Abstract

  We demonstrated that molecular clones of the v-myb oncogene of avian myeloblastosis virus (AMV) can direct the synthesis of p48v-myb both in avian and mammalian cells which are not targets for transformation by AMV. To accomplish this, we constructed dominantly selectable avian leukosis virus derivatives which efficiently coexpress the protein products of the Tn5 neo gene and the v-myb oncogene. The use of chemically transformed QT6 quail cells for proviral DNA transfection or retroviral infection, followed by G418 selection, allowed the generation of cell lines which continuously produce both undeleted infectious neo-myb viral stocks and p48v-myb. The presence of a simian virus 40 origin of replication in the proviral plasmids also permitted high-level transient expression of p48v-myb in simian COS cells without intervening cycles of potentially mutagenic retroviral replication. These experiments establish that the previously reported DNA sequence of v-myb does in fact encode p48v-myb, the transforming protein of AMV.

  View details for Web of Science ID A1986D228300010

  View details for PubMedID 3016296

• ANTIBODIES TO THE EVOLUTIONARILY CONSERVED AMINO-TERMINAL REGION OF THE V-MYB-ENCODED PROTEIN DETECT THE C-MYB PROTEIN IN WIDELY DIVERGENT METAZOAN SPECIES PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Boyle, W. J., Lipsick, J. S., Baluda, M. A. 1986; 83 (13): 4685-4689

  Abstract

  Antibodies directed against a bacterial fusion protein that contains the domain encoded by the highly evolutionarily conserved 5' one-third of the v-myb oncogene of avian myeloblastosis virus (AMV) detect the protein products of various members of the myb gene family. Immunoprecipitation or immunoblot analyses with these antibodies yielded the following information. First, the products of the v-myb oncogenes of AMV (p48v-myb) and of E26 virus (p135gag-myb-ets) contain this highly conserved amino acid sequence, as previously hypothesized. Second, p75c-myb, the product of the chicken c-myb protooncogene, also contains this protein domain. Third, these antibodies have identified the products of the human, murine, and Drosophila c-myb genes, which were all found to be nuclear proteins of Mr 75,000-80,000. The human c-myb protein product is present in immature cells of the erythroid, myeloid, and lymphoid lineages.

  View details for Web of Science ID A1986C992000023

  View details for PubMedID 3088565

• The myb oncogene. Gene amplification and analysis Lipsick, J. S., Baluda, M. A. 1986; 4: 73-98

  Abstract

  The highly conserved, single copy c-myb gene has been independently transduced by two avian acute leukemia viruses, AMV and E26. This gene has also undergone insertional mutagenesis by non-acutely transforming murine leukemia viruses in a number of hematopoietic tumors. The common denominator of these retroviral activations of c-myb appears to be truncation of the normal coding region at either or both ends. The role of point mutations in myb-induced leukemogenesis is currently unknown. The products of the c-myb gene and its altered viral counterparts are nuclear proteins, a large fraction of which are associated with the nuclear matrix. In addition, the myb gene products have short half-lives and bind DNA in vitro. These features suggest that myb may act by regulating DNA replication or transcription. Consistent with this notion, the expression of c-myb is cell cycle dependent in several cell types. However, the abundant expression of c-myb in the thymus is not similarly regulated and may serve a different function. The expression of c-myb appears not to be limited to hematopoietic tissues as previously thought and the nature of the hematopoietic specificity of transformation by v-myb is not currently understood. Nevertheless, hematopoietic growth factors and their receptors appear to play an important role in such transformation. Two new experimental systems for studying myb have recently been described. First, the discovery of a myb-related gene in Drosophila should allow the application of powerful classical and molecular genetic approaches. The functional similarity of this distantly related gene to the much more closely related avian and mammalian myb genes is unknown. Second, recent studies of murine myb in normal and abnormal hematopoiesis offers several advantages relative to the avian system, such as in-bred animal strains, a wealth of specific cell-surface markers, and cloned hematopoietic growth factor and receptor genes. Isolation or construction of an acutely transforming murine myb retrovirus may thus be very useful. Several obvious goals for future research will be to define the function of myb proteins within the nucleus, to understand the regulation of myb expression during the cell cycle, to establish which molecular alterations are essential for converting c-myb into a transforming gene, and the determine the role of myb in human malignancies.

  View details for PubMedID 3333362

• CRYPTOCOCCAL MYOCARDITIS IN ACQUIRED IMMUNE-DEFICIENCY SYNDROME AMERICAN JOURNAL OF CARDIOLOGY Lewis, W., Lipsick, J., CAMMAROSANO, C. 1985; 55 (9): 1240-1240

  View details for Web of Science ID A1985AFS5600032

  View details for PubMedID 3984913

• BIOLOGICALLY-ACTIVE PROVIRAL CLONE OF MYELOBLASTOSIS-ASSOCIATED VIRUS TYPE-1 - IMPLICATIONS FOR THE GENESIS OF AVIAN-MYELOBLASTOSIS VIRUS JOURNAL OF VIROLOGY Perbal, B., Lipsick, J. S., Svoboda, J., Silva, R. F., Baluda, M. A. 1985; 56 (1): 240-244

  Abstract

  A biologically active myeloblastosis-associated virus (MAV) provirus was cloned from a bacteriophage recombinant library constructed from leukemic chicken myeloblast DNA. The restriction endonuclease map of this clone was consistent with that of a type 1 MAV (MAV-1). Interference assays of virus recovered from cultured chicken embryo fibroblasts after DNA transfection established that the provirus was infectious and confirmed that it belonged to avian retrovirus subgroup A (type 1). Antipeptide antibodies raised against the env-encoded carboxyl terminus of p48myb, the transforming protein of avian myeloblastosis virus, specifically immunoprecipitated the gp37env from quail cells transfected with MAV-1 proviral DNA but not from cells infected with MAV-2. This suggests that MAV-1 rather than MAV-2 is the progenitor helper virus from which avian myeloblastosis virus arose by the transduction of cellular proto-oncogene sequences.

  View details for Web of Science ID A1985AQY6700031

  View details for PubMedID 2993653

• AVIAN-MYELOBLASTOSIS VIRUS AND E26 VIRUS ONCOGENE PRODUCTS ARE NUCLEAR PROTEINS PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA-BIOLOGICAL SCIENCES Boyle, W. J., Lampert, M. A., Lipsick, J. S., Baluda, M. A. 1984; 81 (14): 4265-4269

  Abstract

  The defective acute leukemia viruses avian myeloblastosis virus (AMV) and E26 virus each contain an inserted cellular sequence related to the same highly conserved cellular gene, proto-amv. The oncogenes of these two retroviruses differ from this cellular proto-oncogene in gene structure, transcript structure, and gene product. The product of the AMV oncogene (myb) is a 48,000 Mr protein, p48myb, encoded by a transduced segment (amv) of proto-amv flanked by short helper-virus-derived terminal sequences. The E26 virus oncogene product is a 135,000 Mr protein, p135gag-amve-ets, encoded by significant portions of a viral structural gene (gag), sequences related to proto-amv (amve), and additional E26-specific sequences (ets) transduced from cellular proto-ets. Both p48myb and p135gag-amve-ets transforming proteins are located in the nucleus of cells transformed by these viruses. A protein of 110,000 Mr which is specifically immunoprecipitated by antisera to amv peptides and may be the product of the normal cellular gene (proto-amv) has been located in the cytoplasm of cells that express proto-amv mRNA.

  View details for Web of Science ID A1984TD00100006

  View details for PubMedID 6087315

• ONCOGENES - IMPLICATIONS FOR THE DIAGNOSIS AND TREATMENT OF CANCER ANNALS OF INTERNAL MEDICINE Cline, M. J., Slamon, D. J., Lipsick, J. S. 1984; 101 (2): 223-233

  Abstract

  Cellular oncogenes comprise a small family of genes, highly conserved throughout vertebrate evolution, that code for proteins with diverse functions including DNA binding, protein kinase, and cellular growth factor activities. Cellular oncogenes are important in certain aspects of the proliferation and differentiation of normal cells. Under some circumstances these genes may also induce malignant transformation of normal cells. Various mechanisms may underlie their involvement in carcinogenesis. Incorporation of all, or part of, cellular oncogenes into RNA tumor viruses, mutations in gene structure, or translocation of cellular oncogenes from one chromosome to another may all be associated with the induction of malignant change in cells. In some of these situations altered oncogene products are made. Knowledge about the biology of oncogenes may lead to improved techniques for cancer detection and perhaps new approaches to cancer treatment.

  View details for Web of Science ID A1984TE11100017

  View details for PubMedID 6331242

• IDENTIFICATION OF THE LEUKEMOGENIC PROTEIN OF AVIAN-MYELOBLASTOSIS VIRUS AND OF ITS NORMAL CELLULAR HOMOLOG PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA-BIOLOGICAL SCIENCES Boyle, W. J., Lipsick, J. S., Reddy, E. P., Baluda, M. A. 1983; 80 (10): 2834-2838

  Abstract

  The genome of the replication-defective avian myeloblastosis virus (AMV) contains an inserted cellular sequence (amv) that is part of the oncogene responsible for acute myeloblastic leukemia in chickens infected with AMV. Three antisera raised against distinct synthetic peptides predicted from the long open reading frame of amv specifically precipitated the same 48-kilodalton protein (p48amv) from leukemic myeloblasts but not from normal hematopoietic tissue, fibroblasts, or from fibroblasts infected with the AMV helper virus, MAV-1 (myeloblastosis-associated virus type 1). p48amv is not glycosylated or phosphorylated and does not appear to act as a protein kinase in vitro. The same three antisera that recognized p48amv also specifically precipitated a common 110-kilodalton protein from normal uninfected hematopoietic tissue. This normal cellular homologue of the AMV leukemogenic protein, p110proto-amv, was not present in normal fibroblasts, MAV-1 infected fibroblasts, or, interestingly, in some leukemic myeloblasts. We conclude that p48amv is the leukemogenic product of an altered, transduced, partial protooncogene. Short helper-virus sequences provide its carboxyl terminus and also may provide the amino terminus.

  View details for Web of Science ID A1983QR49600006

  View details for PubMedID 6304685

• DIFFERENTIATION AND ACTIVATION OF NU/NU SPLENIC T-CELL PRECURSORS BY MATURE PERIPHERAL T-CELLS IN THE ABSENCE OF THYMUS JOURNAL OF IMMUNOLOGY Lipsick, J. S., SERUNIAN, L., Sato, V. L., KAPLAN, N. O. 1982; 129 (1): 40-45

  Abstract

  Nu/nu splenic T cell precursors lacking significant Thy-1 surface antigen are driven in vitro to proliferate and express Thy-1 during incubation with activated, nondividing peripheral T cells in the absence of thymus or thymic extracts. The precursors are present in nu/+ spleen as well, and are phenotypically similar to thymocyte precursors assayed in vivo. The nondividing inducer cells required are Lyt-2-, Thy-1+ T cells present in nu/+ but not nu/nu spleen and show no MHC restriction in the induction process. Using this in vitro assay, we preliminarily identified a monoclonal rat anti-mouse brain antibody that lyses nu/nu responding T cell presursors in the presence of complement. The ability of mature peripheral T cells to induce the differentiation and activation of T cell precursors in the complete absence of thymus appears to explain the grossly different effects of neonatal and adult thymectomy or thymic involution on immunocompetence. In addition, we showed that the limiting cell in three-cell mitogenic response of normal spleen cells to Con A is likely the macrophage by similar analysis of nondividing inducer cell requirements. This finding allows the assignment of a unique order to this three-cell response.

  View details for Web of Science ID A1982NU21100009

  View details for PubMedID 6177757

• INDUCTION OF LYMPHOMA IN ANTIGENICALLY STIMULATED ATHYMIC MICE CANCER RESEARCH Baird, S. M., Beattie, G. M., LANNOM, R. A., Lipsick, J. S., Jensen, F. C., KAPLAN, N. O. 1982; 42 (1): 198-206

  Abstract

  Athymic mice infected with pinworms or carrying human tumor xenografts frequently develop a lymphoproliferative disorder which eventually leads to lymphoma. By immunofluorescent analysis of involved tissues, the lymphomas appear to be mixtures of null cells, B-cells, and T-cells. When each lymphoma is established in tissue culture, a predominant cell type grows out. We have now established lymphoma lines of null cells, B-cells, and T-cells. Lymphoma development is preceded by the secretion into the bloodstream of large amounts of murine leukemia virus M.W. 70,000 glycoprotein antigen; however, very little virus is produced. In vivo, the expression of viral envelope antigen appears within a few days after human tumor transplantation and precedes the development of lymphoma by about a month. Cells expressing viral antigens are first seen in the diffuse cortex of lymph nodes and the periarteriolar white sheath of the spleen, the tissue domains in which lymphomas also first appear.

  View details for Web of Science ID A1982MW29600034

  View details for PubMedID 7053850

• INTERLEUKIN-2 IS MITOGENIC FOR NU-NU AND NU-+ MURINE SPLEEN-CELLS PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA-BIOLOGICAL SCIENCES Lipsick, J. S., KAPLAN, N. O. 1981; 78 (4): 2398-2402

  Abstract

  Interleukin 2 (IL-2), a product of activated T lymphocytes but not of T-cell-deficient spleen cells of congenitally athymic (nu/nu) mice, is shown to be strongly mitogenic for spleen cells from both nu/nu and nu/+ mice in the absence of serum. This response does not depend on the presence of additional mitogen or antigen as has been previously reported and is dose dependent with respect to IL-2. Plots of the logarithm of cell number versus logarithm of response indicate that IL-2-induced mitogenesis of nu/nu spleen cells is a one-cell event (slope = 1.28 +/- 0.21, mean +/- SD), whereas the response of nu/+ spleen cells to concanavalin A is a three-cell event (slope = 3.18 +/- 0.16). The nu/nu spleen cells proliferating in response to IL-2 are at least 80% T lymphocytes by the third day of culture, as demontrated by lysis with monoclonal anti-Thy-1 antibody and complement. These results suggest that one of the major roles of the thymus may be to induce IL-2 production in a subpopulation of lymphocytes that is then persistent.

  View details for Web of Science ID A1981LP04000077

  View details for PubMedID 6787601

• INDUCTION OF LYMPHOCYTE-T AND LYMPHOCYTE-B RESPONSES IN ANTIGENICALLY STIMULATED ATHYMIC MICE CANCER RESEARCH Beattie, G. M., Baird, S. M., Lipsick, J. S., LANNOM, R. A., KAPLAN, N. O. 1981; 41 (6): 2322-2327

  Abstract

  Antigenic stimulation of athymic mice on the BALB/c background by infection with the pinworms Aspiculuris tetraptera and Syphacia obvelata or by xenografts of human tumors induced a proliferation of T- and B-lymphocytes in spleen and lymph nodes and occasional germinal center formation. The proliferating T-lymphocytes showed greater fluorescence per cell than the Thy 1-positive cells from unstimulated athymic mice when examined by cytofluorography using anti-Thy 1 antiserum. The proliferating T-lymphocytes were shown to be functional by their ability to help mount an in vivo antibody response to sheep erythrocytes and other thymus-dependent antigens. Spleen cells cultures taken from mice at early stages of antigenic stimulation responded in vitro to the thymus-dependent mitogens concanavalin A and phytohemagglutinin. However, spleen cell cultures taken from mice chronically stimulated by foreign antigens were apparently already maximally stimulated and showed no further stimulation when incubated with concanavalin A or phytohemagglutinin in vitro.

  View details for Web of Science ID A1981LS27400046

  View details for PubMedID 6972250

• STREPTOZOTOCIN-INDUCED DIABETES IN ATHYMIC AND CONVENTIONAL BALB-C MICE DIABETES Beattie, G., LANNOM, R., Lipsick, J., KAPLAN, N. O., OSLER, A. G. 1980; 29 (2): 146-150

  Abstract

  Spleen cells from conventional BALB/c or athymic mice with streptozotocin (SZ)-induced hyperglycemia failed to raise blood sugar levels when injected into athymic or thymus-sufficient recipients. Passive transfer efforts were unsuccessful despite variations in donor-recipient pairs with respect to age, thymic function, or time after sensitization of donor mice. Athymic mice develop hyperglycemia following SZ but fail to mount an inflammatory lymphocyte infiltration. In contrast, the heterozygotes show a marked cellular response, which seems to follow the onset of hyperglycemia. The injection of spleen cells from thymus-sufficient mice to athymic recipients confers immunologic competence on the latter as tested by antibody formation to sheep erythrocytes.

  View details for Web of Science ID A1980JN01300011

  View details for PubMedID 6986300

• LECTINS FROM CHICKEN TISSUES ARE MITOGENIC FOR THY-1 NEGATIVE MURINE SPLEEN-CELLS BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS Lipsick, J. S., Beyer, E. C., Barondes, S. H., KAPLAN, N. O. 1980; 97 (1): 56-61

  View details for Web of Science ID A1980KR88100009

  View details for PubMedID 7458941

• PASSIVE TRANSFER OF LYMPHOCYTES FROM DIABETIC MAN TO ATHYMIC MOUSE LANCET Lipsick, J., Beattie, G., OSLER, A. G., KAPLAN, N. O. 1979; 1 (8129): 1290-1291

  View details for Web of Science ID A1979GZ98900021

  View details for PubMedID 87744

• FLAVOKINASE AND FAD SYNTHETASE FROM BACILLUS-SUBTILIS SPECIFIC FOR REDUCED FLAVINS JOURNAL OF BIOLOGICAL CHEMISTRY KEARNEY, E. B., Goldenberg, J., Lipsick, J., Perl, M. 1979; 254 (19): 9551-9557

  Abstract

  A flavokinase preparation from Bacillus subtilis is described which catalyzes the phosphorylation of reduced, but not oxidized, riboflavin. The enzyme is distinguished from other known flavokinases also in having an unusually low Km for the flavin substrate, 50 to 100 nM. ATP is the obligatory phosphate donor; one ATP is utilized for each FMNH2 formed. Mg2+ or Zn2+ is required for the reaction; Co2+ and Mn2+ will substitute, but less effectively. The same enzyme preparation catalyzes the synthesis of FADH2 from FMNH2 and ATP, but not the synthesis of FAD from FMN and ATP. FADH2 is also formed from reduced riboflavin, presumably by sequential flavokinase and FAD synthetase action. Zn2+ cannot replace Mg2+ in FADH2 formation. The reverse reaction, formation of FMN from FAD, occurs only with reduced FAD, giving rise to FMNH2, and is dependent on the presence of inorganic pyrophosphate. The enzyme thus appears to be an FADH2 pyrophosphorylase. The two enzymatic activities, flavokinase and FADH2 pyrophosphorylase, although not separated during the purification procedure, are distinguished by differences in metal ion specificity, in concentration dependence for ATP (apparent Km for ATP = 300 microM for FADH2 synthesis and 6.5 microM for flavokinase), and in the inhibitory effects of riboflavin analogues.

  View details for Web of Science ID A1979HQ50700036

  View details for PubMedID 226520

• TRANSPORT AND BINDING OF RIBOFLAVIN BY BACILLUS-SUBTILIS JOURNAL OF BIOLOGICAL CHEMISTRY Cecchini, G., Perl, M., Lipsick, J., Singer, T. P., KEARNEY, E. B. 1979; 254 (15): 7295-7301

  Abstract

  Riboflavine uptake and membrane-associated riboflavin-binding activity has been investigated in Bacillus subtilis. Riboflavin uptake proceeds via a system whose general properties are indicative of a carrier-mediated process: it is inhibited by substrate analogues, exhibits saturation kinetics, and is temperature-dependent. The organism concentrates riboflavin primarily as the phosphorylated cofactors FMN and FAD. Energy is required for uptake but whether the energy demand is required for both uptake and phosphorylation or only for the phosphorylation step is not known. Membrane-associated binding activity for riboflavin has also been demonstrated in membrane vesicles prepared from B. subtilis, and the binding component can be "solubilized" with Triton X-100. Evidence supporting the function of the binding component in riboflavin uptake by the intact cells includes the following. (i) Riboflavin analogues inhibit binding and uptake to nearly the same extent and with similar specificity of action. (ii) The KD for riboflavin-binding and the Km for uptake are in the same range. Similarly the Ki determined for the inhibitory analogue 5-deazariboflavin in the uptake assay and the KD for its interaction with the riboflavin-binding component of membrane vesicles are in the same range. (iii) Uptake in cells and binding in vesicles vary in the same direction with differences in growth conditions.

  View details for Web of Science ID A1979HG69900075

  View details for PubMedID 110806

• On the Wire (poem) Lipsick, J. The Atlantic. 1974

  Abstract

  Lipsick, J. 1974. On the Wire (poem). Atlantic Monthly 234:61. [https://cdn.theatlantic.com/media/archives/1974/07/234-1/132612162.pdf]